microbiology final
All viruses follow the same general 5 steps when completing their life cycle (i.e. starting with one virus infecting a cell and ending up with multiple viruses getting released). What are these and what is happening at each step?
1) attachment: virus attaches to receptor on host cell - specific 2) penetration: virus enters host cell by direct fusion or endocytosis 3) uncoating: virus sheds capsid and releases genetic information 4) replication: virus takes over host machinery to produce new viral proteins and genetic material 5) release: release of new virus from host cell - lysis of host cell wall or budding
What features of the 16S rRNA gene make it useful for identifying bacteria in the microbial community samples?
1) conservation 2) variability 3) abundance 4) standardization
How do microbes respond to stress? Outline the 4 general steps involved in any stress response.
1) sensing stress: transmembrane proteins sense signal and can be absolute or dynamic 2) signal transduction: chain reaction with protein modification 3) response: gene regulation increased or decreased 4) resolution: return to normal state ones stressor is removed
What are chaperones and why are they needed in some secretion systems? Give two examples of chaperones that play a role in secretion. What do you think would happen if you deleted these factors?
Chaperones promote folding, block aggregation, facilitate protein degregation - they are needed in secretion systems to bind to proteins and prevent them from folding during their transportation through the cell - SecB: binds the protein and prevents it from folding while going through Sec channel -SurA: prevents protein from aggregating , recognizes and binds to outer membrane proteins as they are being synthesized and delivers them to the BAM complex - without these factors proteins would fold prematurely and take different shapes causing them to lose function. they could also degrade without proper folding and wouldn't be able to be transported
Explain how CheA/CheY controls flagellar motion. How does this relate to a run vs tumble? Explain why the rotation direction of the flagella needs to be controlled
CheA sensor gets signal then autophosphorylates and triggers response to CheY which runs down the cell and causes flagellar rotation from default CCW to CW causing a tumble - direction needs to be in control in order for bacteria to steer clear of danger - phosphorylation of CheA then CheY causes tumble to avoid danger from flagella changing direction
Explain, from an evolutionary and geological time perspective, why there are qualities that are unique to protists, and why protists are so diverse as a group.
Evolutionary: protists are some of the earliest eukaryotic organisms so they had time to evolve and adapt leading to their unique features Geological: vast range of environments where they can survive - aquatic/moist, extreme - hot springs, ice caps high level of genetic diversity, can undergo horizontal gene transfer
What would it mean if a viral growth curve is a flat line? Describe how this might occur.
Flat line on viral growth curve would mean that there is no increase in the number of virus over time. This might occur if the virus is unable to replicate within the host cell or if the host cell is resistant to infection
What is one difference between the gut anatomy of many foregut fermenters and many omnivores?
Foregut fermenters: compartment in digestive track called rumen: fermentation chamber where microorganisms and bacteria break down tough plant material Omnivores: no specialized fermentation chamber, rely of bacterial population in large intestine
Almost all bacteria have a heat shock response. Define heat shock and speculate about the temperature that might cause heat shock for a microbe that grows optimally at 10C a microbe that grows optimally at 30C a microbe that grows optimally at 60C
Heat shock response: protection against excessive heat in order for proteins to survive and not be misfolded and denatured. SigmaH increases affinity of RNA polymerase to heat response regulon that produces DNAK to refold proteins above 25-30C 40-45C 70-75C
Speculate why most of the microbial stress responses involve regulation of many genes at once.
In order to survive and adapt to the stressor, microorganisms need to activate a variety of cellular processes and pathways, which often require the coordinated expression of many different genes by coordinating the expression of multiple genes, microbes can ensure that different aspects of their stress response are working in sync it saves energy and at times of stress, microbes need to ration out energy
In the case study we looked at, when JM cultured E. coli in a medium supplemented with both glucose and lactose, E. coli initially utilized glucose rather than lactose. Why?
In the presence of glucose, cells have reduced amounts of the cAMP-CRP (CAP) allosteric regulator for the lac operon
What is an intermediate host vs a definitive host for a parasite? Distinguish between sexual and asexual reproduction.
Intermediate: temporary residence for parasite (asexual) Definitive: final host in life cycle of parasite where it reaches sexual maturity and reproduces sexually Sexual: two parent organisms contribute genetic material to produce offspring genetically different from parents Asexual: single parent produces offspring
We cannot put viruses on the same phylogenetic tree as bacteria, archaea and eukaryotes. Can we put all viruses on a separate, but single viral phylogenetic tree? If yes, explain how. If no, explain why not.
No- viruses do not have a common ancestor and are not considered to be alive since they cannot carry out metabolism, reproduction, or other functions independently of the host cell
There are about 100 bacterial cells in your intestine for every one bacterial cell on your skin. What is one possible reason for this discrepancy? What lifestyle changes might flip this ratio?
One possible reason is the difference in the environments. In the intestine, it is moist, warm, and nutrient rich that is conducive to many bacterial species, where the skin is relatively dry and harsh Hygiene could flip this - dirty hands - more bacteria grows
What is a PFU and how does it compare to CFU?
PFU: plaque forming unit: number of infections viral particles in a sample, measures level of infectivity - count the number of zones where virus-infected cells have been killed main difference is CFU counts number of bacterial or fungal cells, CFU you count number of colonies after sample spread on plate and growth
Compare and contrast prokaryotic vs eukaryotic cells - name 3 fundamental processes that are conserved in all organisms - name 3 fundamental qualities that are unique to eukaryotes (not found in any prokaryotes)
Prokaryotic: smaller, no nucleus, no membrane-bound organelles, have cell wall all organisms 1) DNA replication 2) protein synthesis 3) metabolism eukaryotes 1) membrane-bound organelles 2) cytoskeleton 3) sexual reproduction
What is the Sec system and how are proteins targeted there? Where is the signal sequence encoded? Does it matter if it is at the N-terminus or C-terminus of the gene? Explain why
Sec system: essential secretion of unfolded proteins that is found in most bacteria - N-terminal signal sequence is targeted by SecA that binds and brings it through SEC membrane channel where SecB binds the protein to ensure it doesn't fold while moving through the membrane the signal sequence is encoded at the N terminus end of the protein where the oldest made amino acids are located signal sequences at the N-terminus are more efficient because the Sec machinery recognizes the signal sequence as the protein is being synthesized and the N-terminus location allows for more efficient recognition and targeting
Compare and contrast the various secretion systems. What elements are shared between secretion systems? What elements are different? why do you think there's so many kinds of secretion systems?
Sec: translocation of unfolded proteins across the cytoplasmic membrane TAT: translocate proteins across the membrane: rarely essential, not found in all species BAM: secretion of outer membrane proteins in G- bacteria different types because cells have evolved to survive in the requirements of their environments
Explain how sigma factors work. What does it recognize, and why does this matter?
Sigma factors provide promotor recognition specificity to RNA polymerase holoenzyme, contribute to DNA strand separation, and then dissociate from the core enzyme following transcription initiation
Outline the general pattern of signal transduction systems and two component systems. What is a histidine kinase? Response regulator? How do sensors normally work?
Signal transduction system: a stressor is present and is sensed by a sensor protein on the cell surface that sends signal to response regulator protein to create cell response Two component systems: proteins that sense and respond to environment Histidine kinase: sensor protein that autophosphoylates after sensing stressor and sends signal into cell via phosphorylation response regulator: inside cell and receives signal from histidine kinase via phosphorylation, activating it to bind to target genes and activate or repress them for cell response sensors work: by having extracellular sensing domain that detects change in the environment and a cytoplasmic signaling domain that communicates the signal to downstream components of the signaling pathway - autophosphorylate
Define the characteristics of stress to a microbe. What are some examples?
Stress is a significant change in a microbe's environment that makes growth a challenge. Examples: Nutrition limitation Temperature change Change in pH, pressure, salinity Soap and detergent
Explain how the lac operon is regulated, including the genes lacZ, lacY, lacA, and lacI. How is transcription controlled for each of these genes? You should understand how LacI binds to the operator (when does it bind) and how CAP activates the lac operon in the absence of glucose. What would you expect in the following conditions? Why? 1) high glucose, high lactose 2) high glucose, no lactose 3) low glucose, high lactose 4) low glucose, no lactose
The lac operon in bacteria consists of three structural genes, lacZ, lacY, and lacA, that encode proteins involved in lactose metabolism and a regulatory gene, lacI, that controls the expression of the operon. Transcription of the lac operon is regulated by the interaction of two regulatory proteins, LacI and cAMP receptor protein (CRP or CAP), with specific DNA sequences in the promoter region of the operon. LacI is a repressor protein that binds to the operator sequence of the lac operon and prevents transcription of the structural genes in the absence of lactose. When lactose is present, it is converted to allolactose, which binds to LacI and causes a conformational change that reduces its affinity for the operator, allowing transcription to proceed. CRP/CAP is an activator protein that binds to a specific sequence upstream of the promoter in the presence of cAMP. The level of cAMP in the cell is inversely proportional to the level of glucose. In the absence of glucose, cAMP levels increase, and CRP/CAP binds to the promoter, increasing transcription of the operon. In the presence of both high glucose and high lactose, the cell will preferentially use glucose as the energy source, and the cAMP levels will be low, leading to reduced binding of CRP/CAP to the promoter. The low CRP/CAP activity will result in low levels of transcription of the lac operon, even though lactose is present. In the presence of high glucose and no lactose, LacI will bind to the operator and repress transcription of the operon. The low cAMP levels will also reduce the binding of CRP/CAP, further reducing transcription. In the presence of low glucose and high lactose, LacI repression will be relieved due to the binding of allolactose, allowing transcription of the operon. Additionally, the low glucose levels will lead to increased cAMP levels, which will promote the binding of CRP/CAP to the promoter and further increase transcription. In the absence of both glucose and lactose, the lac operon will be repressed by LacI, and there will be low levels of transcription.
Scientists can alter the DNA in a promoter so that a transcriptional activator becomes a transcriptional repressor. Explain how this could be done.
The sequence that the activator normally binds to is now placed in front of the promoter so now when it is in its active form and binds DNA, it is physically blocking the transcription machinery and represses that operon
Can viruses sense and respond to their environment? Is it possible for viruses to have two component systems? Explain how this could work or why it would not work.
Viruses cannot sense or respond to their environment in the same way bacteria does because they lack the mechanisms to do so, but some may be able to do so in a more basic mechanism by sensing pH. The influenza virus has a two-componant system since it has an outer envelope and an inner core that works together to infect the host cells
What is the benefit of using sigma factors to regulate a regulon?
a set of genes and full pathways can be turned on or off
Bacteria express different things during log phase and stationary phase. What kinds of genes would you expect to be expressed a) mostly during log phase b) mostly during stationary phase c) similarly during both Why
a) genes involved in DNA replication, cell division, metabolic pathways for energy and building blocks since that is the phase of active growth and division b) stress response genes, DNA repair since that is the phase of nutrient depletion where cells are trying to survive c) basic cell processes like transcription and translation
what is the purpose of bacterial endospores
allow a bacterium to survive extended periods of heat or dryness
What distinguishes apicomplexans? Define apical complex, rhoptry, microneme. Are these secretion organelles similar to bacterial secretion systems?
apical complex, complex life cycle - two or more hosts, asexual and sexual replication, plastids apical complex: structure at one end of the cell involved in host cell invasion rhoptry: secretory organelles that secrete proteins into host cell upon invation micronemes: secretory organelles that contain proteins involved in host attachment an penetration share the similarity that secretion of effector proteins that manipulate cell functions and enable infection, some of the proteins share similar structures and functions - may have evolved from common ancestral system
Name two apicomplexan parasites and two kinetoplastid parasites (genus and species)
apicomplexan: 1) babesia microti 2) toxoplasma gondii kinetoplastid: 1) trypanosoma cruzi 2) trypanosoma brucei
The most abundant bacterial species living in the average healthy adult human gut belong to which two phyla?
bacteriodetes firmicutes
what is a virus that enters a bacterial cell
bacteriophage
typically viruses form what around their nucleic acid
capsid
What is chemotaxis? Can bacteria swim directly towards a chemoattractant?
chemotaxis: ability of bacteria to move towards or away from chemical gradients in their environment to navigate towards nutrients and avoid toxins and other harmful compounds bacteria do not swim directly towards a chemoattractant, they do runs and tumbles
What is the defining feature of the kinetoplastids? What is contained in the kinetoplast? (don't need to know the function)
defining feature: presence of a kinetoplast which is a structure within the mitochondrion. - kenetoplast contains thousands of copies of the mitochondrial genome
Compare and contrast prokaryotic and eukaryotic flagella. How do their structures and function differ?
differences: structure: prokaryotic are made of single protein called flagellin in helical structure, eukaryotic made of microtubules in 9+2 pattern function: prokaryotic rotate like propeller to generate movement, eukaryotic whip-like motion similar: involved in movement and use energy
How do bacteria sense a rise in temperature? What is the sensor? What is the response regulator? How do they activate the oH regulon? How do they turn it off?
folded mRNA strands are melted exposing SD that allows for sigmaH to be made in high amounts, causing it to bind to RNA polymerase and turning on heat response genes in heat shock regulon sensor is sigmaH response regulator is DNAK Turned off by FTSH that degrades the bound DNAK and sigmaH and turns off the system
Explain the difference between a gene, operon, and regulon
gene: segment of DNA on a chromosome operon: one or more genes transcribed into a single RNA molecule and under control of a single regulatory site regulon: more than one operon under control of a single regulatory protein
What is one difference between a typical mouse (with gut bacteria) and germ-free mouse? If we engineered a mouse that secreted large amounts of antimicrobial proteins into the intestine, preventing their gut form being colonized by bacteria, would you predict that mouse to do well in the wild?
germ-free mouse has no resident gut bacteria while bacteria can cause disease in the host, they also play important roles in digestion, immunity so mouse could have trouble surviving in the wild - bad immune system, more vulnerable to pathogens in the wild
When an E. coli culture shifts between glucose and lactose utilization, a lag phase is observed. However, when E. coli is shifted form 30C to 40C, the growth rate continues swiftly but no lag is observed. Using your knowledge about how these systems are regulated, explain why.
glucose to lactose: E. coli needs to activate the genes required for lactose utilization which takes time and requires synthesis of new proteins causing the lag phase 30C to 40C: no lag phase: heat shock response is a pre-existing stress response system that is already activated at low levels even at optimal temperature. when temperature increases the heat shock response is rapidly upregulated and chaperones and proteases that are already present are activated rapidly
What is a hemagglutination assay and how does it work? Do you think a similar assay could/would be used to quantify bacteria? Why or why not?
hemagglutination assay: used to detect presence of antibodies or viruses in a sample based on the ability to clump together red blood cells no feasible to quantify bacteria because bacteria wouldn't make red blood cells clump
What is a housekeeping sigma factor vs alternative sigma factor?
housekeeping sigma factors: always on for essential genes alternative sigma factors: turned on periodically for specialized genes
Are there any limitations - what would be the problem if the cell only used sigma factors as the only way to regulate expression?
if the cell only used sigma factors they would need way more proteins and it would waste materials
Draw a graph of a viral replication curve with # of viruses on the y-axis and time on the x-axis - what is each stage called and what is happening at each stage? - If I took the same graph, but told you it was a bacterial growth curve and changed the y-axis to # of bacteria, how would you interpret the data? The lines look similar, does that mean the same things are happening? Explain.
lag - virus prepares for replication, exponential- replication, stationary- replication slows, resources of host cell used up, decline (death) - immune response, death of host cell shape same but different processes- bacteria replicate independently, viruses require hosts - nutrients limit bacteria replication
polasmodium is the cause of what disease
malaria
the basic structure of a virus contains what
nucleic acid, protein coat
What is one problematic aspect of therapeutic fecal microbiota transplantation in humans?
potential for transmission of infectious agents from the donor to the recipient. could also be transferring antibiotic-resistant bacteria or other undesirable microbial traits
what is the infectious substance of prions
protein
In the last few lectures, we've discussed several regulatory systems. Which is the most simple (least parts) to turn on? Which is the most complex? a) lactose utilization b) oH regulon/heat shock response c) stationary phase d) quorum sensing in gram positive
simple: lactose utilization: - only one regulatory protein (LacI) and a single operator sequence most complex: oH regulon/heat shock response: multiple genes and regulatory proteins that work together to respond to heat stress. as well as chaperones and proteases that help protect the proteins from denaturation and promote refolding. multiple regulatory pathways: two-component system
Describe how a small RNA (sRNA) might lead to reduced protein amount in a bacterial cell
small sRNA regulate gene expression by base pairing with mRNA
Understand the different kinds of motility, and how these kinds of movements are achieved. (swimming, swarming, twitching, gliding, sliding)
swimming: smooth straight motion with flagella swarming: groups of rod-shaped cells push each other and move each other with slime layer secreted twitching: running and tumbling in jerky motion of G-, attach to surface of host cells or inanimate objects gliding: glide on solid surface in flaggela-like motion but without flagella sliding: cells pulled away from each other, can adjust their length (extend to attach and then retract to move)
How might you grow a gut bacterial species in the laboratory if it can't tolerate oxygen? What could happen to your gut microbiota if you swallowed a device that continually released oxygen as it passed through your intestines?
use anaerobic chambers, filled with gas with no oxygen, instead nitrogen or hydrogen oxygen damages cell membranes and structures of anaerobic bacteria leading to their death. this would shift the composition of the gut microbiota - sickness of host since many anaerobic bacteria are important for maintaining gut health
What is a virus? What are 3 features present in all viruses (defining features)?
virus: submicroscopic infections agent that can only replicate in the cells of living organisms, have small amount of genetic material, surrounded by capsid, some may have lipid envelope 1) genetic material 2) capsid 3) host dependence
What are viruses made of? Can they have all 4 macromolecules (nucleic acid, protein, lipid, carbohydrate) in the virion? How can you classify viruses based on the components of the virion alone (i.e. without knowing any sequence information or life cycle details)?
viruses made of genetic material, capsid, and in some cases a lipid envelope All viruses have nucleic acid (DNA or RNA), proteins, some have lipid envelope, don't have carbohydrates but some may acquire from host cell during infection Virion: Naked: no lipid envelope Enveloped: lipid envelope Helical: capsid that forms helix around genetic material Icosahedral: capsid that forms a 20-sided shape