Ch 7- Microbial Growth

Are eukaryotic microbes haploid or diploid?

can be either haploid OR diploid

Are prokaryotic microbes haploid or diploid?

haploid ONLY

Do eukaryotic microbes reproduce asexually or sexually?

can do either, asexual and sexual

Do prokaryotic microbes reproduce asexually or sexually?

asexually ONLY

Name three of the asexual reproductive methods of prokaryotes:

1. Binary fission

2. Budding

3. Filamentous

Binary fission

aseuxal reproduction in which a cell separates into two identical daughter cells

Describe the end result of binary fission.

two progeny cells each with

• its own nucleoid

• a complement of other cellular constituents, such as ribosomes and inclusions, which are dispersed throughout the cytoplasm

Baeocytes

small, spherical, reproductive cells produced by some cyanobacteria through multiple fissions

Which genus of bacteria forms multinucleoid filaments that eventually divide to form uninucleoid spores?

Streptomyces

In all kinds of prokaryotes reproductive methods, which features are shared?

• genome must be replicated and segregated to form distinct nucleoids

• each nucleoid and surrounding cytoplasm must become enclosed in own plasma membrane

Which 3 phases does the bacterial cell cycle consist of?

1. period of growth after cell is born

2. chromosome replication and partitioning period

3. cytokinesis = a septum and daughter cells are formed

Which phase of the eukaryotic cell cycle is similar to the 1st phase of the bacterial cell cycle, which involves growth?

G1 phase of eukaryotic cell cycle

Which phase of the eukaryotic cell cycle functionally corresponds to the second phase of the bacterial cell cycle, which involves chromosome replication ad partitioning?

S and mitosis events of M phase of eukaryotic cycle

How is chromosome replication and partitioning different in the eukaryotic v. bacterial cell cycle?

• eukaryotic = S phase and M phase separated by G2

• bacterial = chromosome replication and partitioning occur concurrently

How is the timing of cytokinesis different in bacteria vs. eukaryotes?

• initial events of cytokinesis occur before chromosome replication and partitioning are complete

• some bacteria are able to initiate new rounds of replication b4 first round of cytokinesis and replication are complete

In rapidly growing cultures, the 2nd and 3rd rounds of DNA replication initiate b4 the original cell division is complete. What is the result of this?

daughter cells inherit partially replicated DNA

An increase in ___ triggers replication initation.

cell mass

Each chromosome or plasmid has a single site at which replication starts called the ________.

origin of replication (ori)

What is the site at which replication is completed and where is it located?

terminus

• located opposite the origin

Where do the proteins needed for chromosome replication assemble?

at the origin

Replisome

DNA synthesizing machinery

• large protein complex that copies the DNA double helix to form two daughter chromosomes

In which direction does DNA replication proceed?

bidirectionally from origin

As the chromosome is replicated, what facilitates the separation of the newly replicated chromosomes?

the two origins move toward opposite ends of the cell

What are the three components for the partitioning system for bacterial chromosomes?

• ParA proteins

• ParB proteins

• parS region

Describe the protein-DNA assembly which is called the Partition complex:

ParB binds to parS sites, then additional ParB proteins bind nearby

• covers several thousand base pairs

Where are the partition complexes located?

• one remains at stalk pole of the dividing cell

• other is guided by ParA to opposite pole

Which region of bacterial chromosomes functions like a eukaryotic centromere?

parS sites

Why are the parS sites suitable to direct the segregation of the two daughter chromosomes?

region that is the first part of the chromosome to be duplicated

How is the movement of the newly replicated chromosome of bacterial cells similar to a relay race?

movement of the newly replicated chromosome occurs by passing the partition complex along a series of ParA proteins

Where are the ParA proteins located?

distributed as a gradient in the cell, most abundant at the opposite pole

Cytokinesis vs. Septation

• cytokinesis = formation of 2 daughter cells following cell division

• septation = forming a cross wall between 2 daughter cells during cell division

What are the 4 steps of septation?

1. selection of site for septum

2. assembly of Z-ring

3. assembly of machinery for peptidoglycan synthesis + other cell wall constituents

4. constriction of cell and septum formation

Which cytoskeletal protein makes up the Z ring?

FtsZ

Which system and proteins determines where the Z-ring will form? How does it work?

Min system

• MinC

• MinD

• MinE

Prevent polymerization of FtsZ in wrong places

• Min C = inhibitor and accumulates near poles, concentrating FtsZ filaments at midcell

Which system oscillates between the poles to prevent FtsZ polymerization at the poles?

Min system

What happens if Z-ring formation is done too early?

Z-ring could constrict around replicating chromosome = prevents proper partitioning

What is Nucleoid occlusion?

mechanism used by many bacteria to prevent premature formation of a septum during binary fission

• chromosome movement coordinated with cell separation

How does nucleoid occlusion prevent the Z ring from forming until the replicated chromosomes have almost completed partitioning to the poles?

• SlmA protein coats chromosome

  • except at termination region

• inhibits FtsZ polymerization

How is the Z-ring attached to the membrane?

indirectly attached through interactions with several adaptor proteins

Treadmilling

individual filaments within Z-ring move around interior circumference of membrane

• results in removal of individual FtsZ monomers from one end of a filament and repolymerization of new monomers at other end

Which interaction with the Z-ring stabilizes the dynamic Z-ring and prevents constriction until two new chromosomes have moved away from midcell?

connection to ter region of chromosome

Divisome

collection of proteins that aggregate at the region in a dividing microbial cell where a septum will form

What is the scaffold upon which enzymes that synthesize the layers of the cell wall assemble?

Z-ring

Bactoprenol

lipid carrier molecule used to carry peptidoglyacn precursors from cytoplasm to periplasm

Steps of peptidoglycan synthesis during bacterial septation

1. synthesis of NAG-NAM-pentapeptide building block, completed in cytoplasm while attached to bactoprenol

2. Carrier-bound building block flipped across protein called MurJ

3. NAG-NAM-pentapeptide released into periplasmic space and inserted into peptidoglycan strand by enzymes (GTases)

Which enzymes crosslink the peptidoglycan strands?

transpeptidases (TPases)

Which enzymes are also called penicillin-binding proteins and why?

Transpeptidases (TPases)

• discovered as molecules that physically associated with penicillin

Which enzyme inserts the NAG-NAM-pentapeptide into a peptidoglycan strand?

glycotransferases (GTases)

Elongasome

complex of enzymes in rods (helps maintain shape) that synthesizes peptidoglycan during cell growth

How does the elongasome differ from the Z-ring?

its scaffold is MreB instead of FtsZ and Z-ring

Which cytoskeletal protein directs the assembly of elongasome proteins like GTases and TPases on periplasmic face of plasma membrane?

MreB

What happens to rod shaped cells when MreB is depleted?

assume a spherical shape

Cocci lack proteins in which family?

MreB

What is the general purpose of the Miin system and nucleoid occlusion?

spatial controls which help place the Z-ring in the cell center

Batch culture

growth of microorganisms in a closed culture vessel without adding fresh or removing old/spent medium

• nutrients consumed, concentrations decline, and wastes accumulate

Function of autolysins in synthesis of new peptidoglycan

autolysins break glycosidic bonds at point of new synthesis of peptidoglycan

Function of bactoprenol in synthesis of new peptidoglycan

hydrophobic lipid that transports precursors of peptidoglycan across membrane

Penicillin, vancomycin, bacitracin

antibiotics that stop bacteria from building their cell walls

Which antibiotic blocks the final cross linking step of peptidoglycan synthesis by binding to transpeptidase enzymes?

Penicillin

What is plotted on the Growth curve?

Logarithm of cell number vs. Time

What are the 5 phases of the Growth curve?

1. Lag

2. Log/exponential

3. Stationary

4. Death

5. Long-term stationary

Which phase of the growth curve refers to the period after the introduction of microbes into fresh culture medium, when there is no increase in cell numbers or mass during batch culture?

Lag phase (1)

Is the lag phase a time of inactivity?

No, cells are synthesizing new components to be able to begin growth and have necessary enzymes for the new medium they’re in

During which 3 scenarios will a lag phase occur?

1. From stationary or death phase to fresh medium

2. From exponential phase to fresh medium of different chemical composition

3. From rich to poor culture (medium shift)

When does a lag phase NOT occur?

From cells growing exponentially to a fresh medium of same chemical composition

During which phase of the growth curve is the microbial population growing at a constant and maximum rate, dividing and doubling at regular intervals?

Exponential phase

During which phase of the growth curve is the population most uniform, in terms of chemical and physiological properties?

Exponential phase

During which phase of the growth curve is the population healthiest?

exponential phase

Which grow faster, prokaryotes or eukaryotes?

prokaryotes

Which grow faster, smaller or larger cells?

smaller

How does the growth rate relate to nutrient concentration?

increases with nutrient concentration, but saturates at sufficiently high nutrient levels

In which phase of the growth curve does population growth cease and the growth curve levels off (horizontal)?

stationary phase

During which phase of the growth curve does population growth cease and waste accumulates?

stationary phase

During which phase of the growth curve does the total number of viable cells remain constant?

stationary phase

Which phase of the growth curve do microbes enter because of nutrient limitation?

stationary phase

What are three reasons microbes may enter the stationary phase?

1. nutrient limitation

2. accumulation of waste products

3. critical population level reached

Why does DnaA protein become less active in stationary phase?

it is the protein that binds to the chromosome’s origin to initiate replication, no further initiation for replication occurs

• conserves the cell’s energy to focus it on processes essential to survival

Can cells growing in batch culture (closed system) remain in stationary phase indefinitely?

no, eventually enter death phase

Describe the death phase of the growth curve

cell death occurs at constant rate =

number of viable cells declines exponentially

What causes death phase?

nutrient deprivation and buildup of toxic wastes causes irreparable harm to the cells

What are the two hypotheses relating to death phase of the growth curve?

1. Viable but not culturable (VBNC)

2. Programmed cell death (PCD)

Which bacteria can be viable but not culturable (VBNC)?

• Escherichia coli

• Vibrio cholerae

• Listeria monocytogenes

What is the long-term stationary phase of the growth curve?

phase after death phase in which population size remain at more/less constant low level, for extended period of time

How long can the long-term stationary phase last?

months to years

How can natural selection be seen in the long-term stationary phase?

bacterial population continually evolves

• actively reproducing cells = those best able to use nutrients released by dying cells and those most tolerant to accumulated toxins

In which phase of the growth curve are birth and death rates balanced?

long-term stationary phase

What is the GASP phenotype, seen in the long-term stationary phase?

Growth advantage in stationary phase

• evolutionary survival mechanism where aged bacterial cells outcompete younger cells

• mutated "GASP" cells can utilize nutrients released by dying cells.

Microbial succession in milk spoilage:

1. lactic acid bacteria

2. fungi

3. proteolytic bacteria

What does a larger growth rate constant (k) indicate?

larger k = faster growth

What does the growth rate constant (K) refer to?

generations per hour

The quantitative aspects of growth are only valid for which cultures/micribes?

those undergoing the exponential phase of binary fission

• only exponential phase has a population acting in synchrony

What is the term for the specific length of time required for a microbial population to double in number?

Generation (doubling) time (g)

A linear log plot of number of cells and time indicates what?

the population is in exponential phase

• blue = log scale

• red = linear scale

Equation for population at time t, for populations reproducing by binary fission:

N_t = N₀ × 2ⁿ

N_t

population at time t

N₀

initial population number

n

number of generations in time t

Equation for n, number of generations + how is it derived?

n = log N_t - long N₀ / 0.301

• N_t = N₀ × 2ⁿ

• log N_t = log N₀ × n*log 2

• n = log N_t - log N₀ / log 2

• n = log N_t - log N₀ / 0.301

log 2 = ?

0.301

Equation for growth rate constant (K) + how is it derived?

k = n/t = log N_t - log N₀ / 0.301t

• n = log N_t - log N₀ / 0.301

• divide by t = log N_t - log N₀ / 0.301t