Topic 4 Flashcards
(114 cards)
1
Q
Most microbial cells divide by
A
binary fission: DNA replicated, cell grows, cytoplasm evenly divided, producing two ~identical daughter cells
2
Q
What happens after replication in binary fission?
A
segregation: DNA to opposite sides of the cell so that each daughter gets one copy
3
Q
What happens after segregation?
A
cells elongate and a septum begins to form to separate the cell into two
once this is complete (formation of cell walls causes cell separation), the division site constricts and the cells are pinched off from one another
4
Q
DNA replication is controlled by
A
binding of protein DnaA to oriC
5
Q
ONE important mechanism for controlling binding of DnaA to oriC
A
via a protein called SeqA (competes with DnaA for oriC binding site)
6
Q
For some bacteria, generation time is shorter than amount of time it takes to replicate chromosome. How ???
A
multiple DNA replication forks can be active at once; essentially, OriC region starts getting replicated again before the first fork is finished
-> occurs in E. coli and Bacillus subtilis
7
Q
Divisomes
A
multi-protein complexes that help identify cell centers to build a septum
8
Q
FtsZ
A
- acts as a divisome; a central protein
- polymerizes to form a ring around circumference of cell at the mid-cell where division will occur
- constricts to help division occur
- recruits proteins to help build cell envelope (SEPTUM)
- depolymerizes at the end
9
Q
nucleoid occlusion
A
when chromosome is at mid-cell, prevents divisome from forming there!
-> push nucleoid to the side to resolve this!
10
Q
Factor in finding the mid-cell besides occlusion
A
FtsZ (Z-ring) inhibitors
- clustered at cell pole and lowest concentration at mid-cell
- EX: “min” system - MinC/MinD at poles
11
Q
T or F. Cell wall must be remodelled but remain intact during cell division to prevent cell from bursting
A
T
12
Q
Cell wall growth - peptidoglycan precursors brought across cytoplasmic membrane by
A
bactoprenol (hydrophobic - lipid)
13
Q
Autolysin
A
break glycolytic bonds for insertion of new monomers for cell wall growth
14
Q
Cell wall growth: transglycosylases and transpeptidase
A
Transglycosylase enzymes form new bonds in sugar backbone; new beta-1,4- linkage
transpeptidase enzymes form new peptide crosslinks; seal up peptides (built by D-alanine)
15
Q
What dictates shape?
A
How cell wall is built during cell growth
16
Q
Rod vs Spherical shaped cells
A
Rod - actin-like cytoskeleton protein MreB ensures that new cell wall is added along its long axis – rod elongates
Spherical - lack MreB; new cell wall added only at midcell; spherical shape is default
17
Q
Alternatives to binary fission (assymetrical)
A
- budding division: new cells frow from old cells by budding off from a particular site on mother cell
- multiple fission: new cells bud off from long extensions called hyphae and then some instances occurs where long hyphal filaments will form multiple septa at once and break off into many new cells
- sometimes cells grow only from one poly to produce cells built from new and old material
18
Q
Growth or culture media
A
(medium, singular) can be highly variable depending on the microbe… and for a given microbe
19
Q
Defined media
A
media prepared by adding precise/known quantities of chemicals to water; know the exact composition
20
Q
Complex media
A
contain extracts or digested organic material with an unknown composition (eg: yeast extract, casein (milk protein) digests)
21
Q
Advantage of defined medium
A
you know what you’re working with
22
Q
The more common type of media
A
Complex media; cheaper, easier, work for a broader array of different microbes
23
Q
Some microbes can make most or all of the organic molecules they need
A
- many microbes that live in nutrient-poor environments
- certain ‘flexible’ microbes that adapt to many different environments (such as E. coli)
24
Q
Other microbes can require a great number of growth factors – things like vitamins, amino acids, purine/pyrimidines, etc.
A
- many organisms have an obligate symbiotic lifestyle
- organisms that live in nutrient-rich environments (such as many lactic acid bacteria)
25
Auxotrophy vs Prototrophy
A - inability to produce a molecule you need for your growth
P - opposite; can produce molecule
26
Media used to isolate out a specific microbe or determine which or if a species of microbe is present
Selective, differential, enrichment
27
Selective media
used to isolate a limited range of microbes (often bacteria) --- this could be a single species; often a combination of positive (nutrients few organisms can grown on) and negative (substances that kill most microbes) selection
28
Differential media
contain some sort of an indicator (eg. a dye that changes colour) when particular organisms are present
29
Enrichment culture
similar idea to selective media ... but less selective and richer medium; promotes growth -- increase numbers from isolates to make it easier to isolate a particular microbe; usually somewhat selective
30
Solid vs Liquid cultures
Solid - useful for isolating single colonies; can identify contamination
Liquid - should generally be started using isolated single cultures streaked on agar plates; came from single colonies on plates
31
The great plate count anomaly
When 'natural' samples are plated on genetic growth plates (mean to be 'non-selective') ... 99.9% of microbes don't grow.
32
T or F. We cannot culture the vast majority of microbes
T! One big reason is syntrophy (microbes feeding off one another =; microbes require other community members to grow)
33
Counting microbial cell numbers
1. Direct count: cells in the sample are counted using a microscope
2. Visible plate counts: a small sample of cells are spread on agar plates and incubated -- colonies counted
3. Turbidimetric: absorbance of light by a microbe growing in a liquid culture is measured in a spec
4. other indirect methods: O2 consumption, CO2 production, metabolic activity; quantitative PCR to determine the number of 'genome equivalents'
34
Much more convenient than colony counting (no dilutions, just OD measurement
Turbidity measurement - creates a standard curve, linear range, etc
35
Significant advantage of turbidity measurements
- not labor intensive
- growth can be continuously or regularly measured over time.
- growth can be visually observed ; "cloudiness"
36
Microbial "growth"
refers to increase in population size -- cell division resulting in multiplying in numbers
37
Generation time
AKA doubling time (exponential phase) - amount of time it takes for one cell to become two (double in numbers and mass); varies greatly depending on microbe and growth conditions
38
Batch cultures
cultures in a fixed volume in a closed (closed system) container like a flask or a test tube; finite amount of time - reach a saturation point where cells cannot grow anymore
39
Continuous cultures
cultures within systems where waste product are being removed and new media fed in; environment can be held constant for a long period of time to allow for extended cultures to take place
40
In _____ cultures, microbes typically exhibit a growth curve that reflects the different stages of growth in this environment
batch
41
Lag phase
period of slow (or no) growth as microbes adjust after they are inoculated into this new environment
42
Exponential phase
growing population doubles at regular intervals; nutrients not yet exhausted, waste products not slowing growth
43
Stationary phase
nutrients begin to be exhausted (limiting) and/or accumulating waste products inhibit growth; little to no net growth (still some growth and death)
44
Decline phase
if you leave cultures long enough, cells will start to die (net decline in cell numbers)... this phase often isn't all relevant
45
Phase used for studies; bacteria are all in a physiologically identical phase
Exponential phase
46
Generation times during exponential phase
- cell numbers double at some regular interval
- generation time can range form less than 20 mins to weeks or more
- generation time = growth time/number of generations
47
Cells can be grown indefinitely in the lab if we can...
remove waste products (and cells) then add fresh media
48
Chemostat
continuous culture device; can be used to grow cells at a steady state; culture volume, number of cells, nutrient-waste status all kept constant
49
Dilution rate
diluting rate (culture volume is steady); volume always added and removed at same rate; this controls growth rate
50
T or F. The larger the dilution rate, the faster the growth
T
51
Planktonic vs Sessile growth
planktonic = free-living organisms in liquid
sessile = growth attached to a surface
52
Growth on surface can develop into
biofilms (cell encased in polysaccharide matrix attached to surface)
53
How can biofilm formation start?
- a planktonic cell can attach to a surface via appendages such as pili, fimbriae or even the flagellum
- colonization begins and the cells grow (multiply) and produce extracellular polysaccharides
- during development, cells change their biological program (express unique combinations/amounts of genes to facilitate this biofilm lifestyle)
- ultimately, some cells will begin to disperse (resume planktonic state ... maybe form a biofilm elsewhere)
54
Psychrophile
optimal growth <15 C; many are killed when exposed to moderate temps
55
Mesophile
most commonly studied and widespread; growth at intermediate temperatures (roughly the same temperatures as us)
56
Thermophile
optimal growth rate <45 C
57
Hyperthermophile
optimal growth rate >80 C
58
Psychotolerant
organisms that can grow at low temps but optimally mesophiles (20-40 C)
59
Acidophiles
microbes that can survive under highly acidic conditions; prefer this
also...acidotolerant (but prefer neutral pH)
60
Alkaliphiles
pH >8
61
Although cytoplasmic pH will vary somewhat in alkaliphiles and acidophiles,
cytoplasmic pH is generally maintained close to neutrality (6-8)
62
Strategies used to maintain pH close to neutral in acidic/alkaline environments
keeping out protons or pumping protons out, creating buffer molecules, etc.
63
____ _________ of enteric pathogens has a big influence on _________ ____ (?)
acid tolerance; infectious dose (minimum number of organisms consumed to become sick)
64
Most significant osmotic factor in nature
Salt concentration
65
Halophiles
live in high salt environments
66
Non-halotolerant but will still produce compatible solutes to help adjust to osmolarity of environment
E. coli
67
Compatible solutes
solutes produced to grow in high solute environments by producing high concentrations of solutes (increase cytoplasmic solute concentration); do not interfere with cell's biochemical processes
68
T or F. Compatible solutes are highly water soluble and inert
T!
69
Microaerophillic bacteria excel in
low O2 environments
70
There are many anoxic environments on the planet:
sediments, bogs, marshes, subsurface, animal intestinal tracts (microaerobic)
71
Enzymes that destroy toxic oxygen species
Catalase, peroxidase, superoxide dismutase, superoxide dismutase/catalase in combination, superoxide reductase
72
Decontamination
neutralization or removal of microbes
73
Disinfection
eliminating or reducing harmful organisms
74
Sterilization
process of killin all microorganisms; if something is sterile, it is completely free of microbes; binary (it is or it isn't)
75
Decimal reduction time
amount of time it takes to reduce number of microbes by a factor of 10 (a log); under a specific set of conditions
76
Thermal death time
amount of time it takes to kill all cells at a given temperature; depends on number of cells (takes longer if a lot of cells)
77
Decimal reduction rat and thermal death time depend on a number of factors:
pH, salt concentration, moisture levels, presence of fats/sugars/proteins in samples (can decrease heat penetration), etc.
78
Common method to prevent spoilage and protect against food-borne pathogens with certain foods/beverages
Pasteurization ; heated for a specific amount of time to eliminate pathogens (5-6 logs), and to reduce spoiling agents
79
Pasteurization for milk
71 C for 15 seconds is common (balance between minimal loss of food quality and safety)
80
In milk, heat-resistant lactic acid bacteria survive _________ and are what cause the milk to spoil over time
pasteurization; most gram neg are killed but some heat-resistant gram pos and lactic acid bacteria will survive
81
This damages DNA and is lethal to microbes at high enough intensities (another method used to sterilize)
UV light; useful for sterilizing SURFACES ; poor penetrating power
82
Improved penetrating power compared to UV light
Ionizing radiation (such as gamma rays); regularly used for sterilizing surgical supplies, labware, and even food
83
Another way to sterilize without using heat (can cause damage to proteins + other molecules)
Filter sterilization; pass liquids through filters with a pore size of ~0.2 microns ; not as reliable as autoclaving
84
Products that reduce microbial numbers are
non-specific mechanisms (harmful to human cells, particularly if ingested) - membrane disruption, protein denaturation, oxidizing agents
85
Sterilants
kill all microbes - formaldehyde
86
Disinfectants
largely for surfaces, kill many/most microbes but not all (not endospores) - lysol
87
Sanitizers
unlike disinfectants and sterilants, less harsh to humans, but generally also less effective - soaps
88
Antiseptics
kill or inhibit growth of microbes, non-toxic enough to use on tissues (external such as wounds) - ethanol
89
Antimicrobial agents
chemical that kills or inhibits the growth of microbes
90
Bacteriolytic (antimicrobial agent)
agents that not only kill cells, but cause them to lyse (no dead cells observed)
91
MIC of a compound against a particular microbe
minimal inhibitory concentration - lowest concentration of a compound that fully inhibits microbial growth
92
How to determine MIC
serial dilutions of the compound ( usually 2x dilutions) - measuring microbial growth
93
Besides MIC, these measure toxicity of chemicals through solid media
Kirby-Bauer test; disk placed on agar plate and it diffuses; closer to disk = higher concentration of microbial agent ; size of zone of inhibition (area with no microbial growth) indicates susceptibility to compound
94
Antibiotics
use specific mechanisms to target microbes, rather than generic and non-specific toxicities such as widely inactivating proteins or disrupting membranes
95
Antibiotics manufactured into creams/ointments that are applied to skin
Topical antibiotics ; others are ingested and must not harm human cells
96
Alexander Fleming
- discovered lysozyme
| - penicillin (first true antibiotic); before it was arsenic compounds o treat syphilis which was toxic!
97
First true antibiotics used in the clinic
Sulfa drugs
98
First effective, ingested antibiotic used in clinics
protonsil (penicillin not used after more than a decade of discovery)
99
This is not effective in test tube, only in live animals
protonsil - a pro-drug that must be processed by the body to active form
100
How do sulfa drugs, specifically protonsil work?
through competitive inhibition; inhibit PABA (foliate biosynthesis
101
Polymyxins
permeabilize outer membrane of gram neg; then disrupt cytoplasmic membrane too ; can use polymyxins in conjunction with another drug (destruct OM and let other drug in)
102
How does penicillin work?
binds to and inhibits activity of PBPs which are proteins that catalyze transpeptidation reaction that crosslink the cell wall
103
T or F. Most successful antibiotics have been isolated from microbes
T! they secrete antibiotics as a competition mechanism
104
Certain Actinomycetes have been a very rich source of antibiotics
Streptomyces - soil-dwelling bacteria that produce large numbers of antibitotics as they have large genomes, many interesting biosynthetic gene clusters, etc.
**Challenge : turning on all biosynthetic clusters
105
T or F. Antibiotics are not just for bacteria
T! Some target eukaryotes such as fungal pathogens but since they have overlapping cell biology with humans, this limits targets and some are only applied topically
106
Polyene antibiotics
- from Streptomyces
- inhibit ergosterol in fungus
- permeabilize membrane and causes cell death
107
How can antibiotic resistance occur?
due to mutation, but more commonly by acquiring antibiotic-resistance genes via horizontal gene transfer
-> spurred on by improper use of antibiotics, over-prescription, extensive use in agriculture, etc.
108
4 mechanisms of antibiotic resistance:
1. Modification of drug target
2. Enzymatic inactivation of the drug
3. Removal from cell by efflux pumps
4. Metabolic bypass
109
Beta-lactamase
antibiotic-resistance gene; enzyme that degrades penicillin
110
Antibiotic derivatives
- improved activity
- more borad spectrum
- less toxicity in humans
- overcome known resistance mechanisms
111
Persistence
when antibiotic-sensitive bacteria includes rare cells that transiently tolerant to antibiotics
112
Dormancy
slowing or shutting off metabolism is a common route to persistence
113
Unlike resistant bacteria, persisters are...
genetically unchanged - can be killed by antibiotics once again after they emerge form their tolerant state
114
Most antibiotics depend on (2) in order to be effective
metabolic activity and often growth (ex: if you're not building a cell wall, inhibiting cell wall synthesis is not effective)
