An increase in population of microbes not not an increase in their size
microbal growth
bacterial cells must ___ to increase their population
divide
bacterial cell division requires
macromolecules ( dna, proteins, lipids and carbs
how many copies of dna are require for each cell to divide
2
one parental cell splits into two equal daughter cells this process is called
binary fission
ori
origin of replication where sequencing begins
ter
termination sequence which signals end of replication
two replisomes ( replication fork with proteins attached) move
bidirectionally ( both directions) from the ori and will eventually meet at ter sequences
ori is attached to the
membrane
ori is attached to the membrane and as the cell elongates, it
pulls the DNA apart
Dna can start the next round of replication before
its done with the first making it a very rapid process
in bacterial cells, each cell continues to replicated its dna resulting in rapidly dividing cells having
more than one copy of dna
generation time can be measured in
minutes
ecoli has a generation time of
30 min
endospores generation time can take
years centuries, etc.
some species of bacteria divide the availability of
nutrients
bacteria do not have a very good supply of nutrients and will not be able to make the
macronutrients they need
___ has a key role in determining the shape and arrangement of cocci
spatial orientation of septation
arrangements of cooli progeny cells depend on
how and where the cell wall forms
how and where the cell wall forms
septation orientation
in the arrangement of cocci, if its ____ then chains are formed ex. streptococci
parallel planes
in the arrangement of cocci, if they are formed in _____ then grapelike clusters are formed. ex. staphyloccocus
random planes
in the arrangement of cocci perpendicular planes form
tetrads and sarcinae.
before the septum forms, the ____ protein forms a ring around the center of the cell
FtsZ
FtsZ is related to ___ a component of cytoskeleton
tubulin
FtsZ requires ____ energy
GTP
a FTS protein is an anchor that connects the FtsZ ring to the cytoplasmic membrane
Zip A
connects FtsZ ring to the membrane and recruits other divisome proteins such as Ftsl and Ftsk
FTS A
FTS proteins are _____ and a cell cant divide and appear as long filamentous strands
nonfunctional
FTS proteins are ______ meaning they are all known bacterial cells
universally distributed
control of FTS proteins is dependent on the
formation of the FtsZ ring
the septum is made after
replication
what does FtsZ protein stand for?
filamenting temperature sensitive mutant Z
are temp sensitive mutants, may be permissive or non permissive temps
FtsZ proteins
temperature which the FtsZ protein has a normal function
permissive temperature
a mutation can be seen in the phenotype of FtsZ protein
non-permissive temperature
temp sensitive mutants are called ___ because in one condition they look normal and in another they dont
conditional mutants
only way to study loss of mutation in FtsZ proteins is do so as a ___
conditional mutant
cell division of FtsZ proteins can be studied by
green fluorescent protein ( GFP)
cinnamaldehyde can be used as an
anti microbial
MreB a cytoskeletal protein is key in determining
cell shape. it elongates the cell
if MreB is deleted from ecoli it will appear as
staphylococcus b.c it wont elongate
makes bacteria a curved shape. It binds one side of the bacterial cell while MreB elongates the cell
crescentin
FtsZ forms a ___ in the middle of the cell
ring
cells that lack mreB gene appear to be
circular
MreB cytoskeletal proteins are located at
rings
why is building and elongating the cell wall in bacteria complicated?
small section of peptidoglycan cell wall have to be broken first and then filled in with a larger piece
are glycoylases that break the glycosidic ( sugar-sugar) bonds between N-acetyl
N-acetylmuramic acid
and N-acetyl glucosamine along
the existing peptidoglycan
Chinese character look" upon gram staining whereby bacterial cells appear in a scattered
pattern. This is a characteristic of
Corynebacteria diphtheria.
bonds between N-acetylmuramic acid
and N-acetyl glucosamine are the bonds that keep ___ in line
NAM-NAG-NAM-NAG
Glycosylases
known as autolysins break the
glycosidic (sugar-sugar) bonds
between N-acetylmuramic acid
and N-acetyl glucosamine along
the existing peptidoglycan.
These breaks disrupt
the cross links in the region
new peptidoglycan monomers with their pentapeptide side groups and their sugars ( NAM-NAG) attach to a nonpolar carrier molecule called_____ which transports them across the cell membrane
bacterophenol
bacterophenol are assembled in the _______ and have to be transported to the _____ layer
cytoplasm, peptidoglycan layer
new peptidoglycan monomers with their pentapeptide side groups and their sugars ( NAM-NAG) attach to a nonpolar carrier molecule called bacterophenol. where they are inserted into the growing peptidoglycan chains. once there, _____ occurs by 2 strands whe
transpeptidation ( cross-linking)
penicillin inhibits the enzyme responsible for _____ and therefore interferes will cell wall synthesis in growing cells
transpeptidation
cell division in bacterial cells results with what kind of poles?
a new pole and a old pole
when bacterial cells divide ____ poles arise
new
with continuous division in a bacterial cell, the poles of each daughter cell differ ___ from one another
chemically
what are the two types of culture media
liquid or broth media and solid media
what kind of culture media is useful in studying growth of a pure culture, also produces bacteria that is uniform in population and description. Useful for freezing or preserving culture
liquid or broth media
gelled culture media, useful for separating mixed cultures from clinical specimens or natural environment . Colonies can be isolated and then isolated colonies can be cultured in a flask with liquid to mass produce it as a pure colony
solid media
progeny of single cell ( all cells of that colony are identical )
colony
chemically defined media and the molar concentrations of all the same elements in the media are kow
defined media
there is an unknown variable and can also provide added micronutrients & growth factors richer than defined media
complex media
a type of media that contains additives
enriched
a media that species will grow however they differ from each other. it allows researchers to identify different types of bacteria their working with
differential media
a media where only target organisms grows therefore only one specimen grows
selective media
composed of ingredients such as peptones and extracts which may very in their chemical composition
complex
composed of precise mixtures of pure chemicals such as ammonium sulfate
chemically defined medium
medium to which additional ingredients have been added that inhibit the growth of many organisms other than the one being sought
slective medium
medium that contains an ingredient that can be changed by certain bacteria in a recognizable way
differential medium
complex medium used routinely in clinical labs. not selective, differential because colonies of hemolytic organism are surrounded by a zone clearing of the Red BC's
blood agar
complex medium used to culture fastidious bacteria, particularly those found in clinical specimens. not selective or differential
chocolate agar
chemically defined medium. used in laboratory experiments to study nutritional requirements of bacteria, not selective or differential
glucose-salts
complex medium used to isolate gram neg rods that typically reside in the intestine. selective b/c bile salts and dyes inhibit gram positive organism and gram neg cocci., differential bc the ph indicator turns red when the sugar in the medium, lactose is
MacConkey agar
complex medium used for routine laboratory work. support the growth of a variety of nonfastidious bacteria
nutrient agar
complex medium used to isolate neisseria species, which are fastidious. selection because it contains antibiotics that inhibit most organisms except neisseria species
thayer martin
can be seen with a scanning electron microscope , grows only withins the cytoplasm of eukaryotic cells. impossible to grow this bacteria without its host cells
reickettsa prowazekii
how to know how many bacteria are present in a sample
plate counting and serial dilutions
describe the process of serial solutions
1 ml is taken from unknown quantity of colony forming units in a flask
transferred into 9 ml of media in a test tube
test tube is vortexed and 1 mil is taken from this tube to another containing 9 ml of media
done several times, diluting concentration of
describe this example of dilution factor 1:10 X 5 = 10^-5
1:10 ( label of 1st test tube, the 2nd would be 1:100), 5 (5th test tube)
Number of bacteria present: 27 colonies x 105 = 2.7 x 106 CFU/ml
When counting plates, accuracy depends on the number of
colony forming units (CFU) that
appear on the plate
when counting plates, Too few result in discrepancies and too many could result in human error
since some colonies may be growing on top of each other and may be counted as
one
colony forming units are therefore considered valid.
30-300
Serial 10-fold Dilutions are commonly used in
counting plates
If there are 5 test
tubes, then it was diluted 10-5 which means to calculate the number of bacteria present multiply
the number of colonies by 105
, likewise, if there were 4 test tubes then they were diluted 104
which means to calculate the number of bac
example of Serial 10-fold Dilutions
The CFU/mL is calculated by the
equation: dilution factor (e.g. 104
) x # of CFU x 10.
Colony forming units (CFU) are
live bacteria that can form colonies
Disadvantage of serial dilutions
time consuming
some cells may be live but did not form colonies and are therefore not counted
advantage of serial dilutions
method is that it is fairly accurate and gives VIABLE or live counts
Cells can also be
counted directly and quickly.
can be used to count cells directly and quickly using a microscope this is called a
Petroff-Hausser Counting Chamber
It has a 25 large squares (1 mm2 x 1/50 mm deep) chamber. Cells from a small
square (0.0025mm2 with depth 0.2mm and volume 0.0005�L) within a larger square is used
when counting.
If for example 10 cells were counted in a small square, then the cells/squar
example of petroff-hausser counting chamber
advantages of Petroff-Hausser
it is easy and quick since no culture is needed
disadvantages of Petroff-Hausser
might get a false high number since dead cells can be counted and
counting errors due to small cells and clumps
it needs 10^6
cells/mL or more to see and calculate accurately
motile cells are very hard to count once
they are moving.
living and dead cells can be distinguished from each other by
fluorescence microscopy
may also be used to count cells (shown in A.). A laser
beam is used and the sample stream is one cell at a time. The cells are counted and can even
be sorted based on their characteristics such as non-GFP producing cells versus GFPproducing
cells as seen
A Flow Cytometer or Coulter Counter
Turbidity or absorbance of microbial growth can be measured using a
spectrophotometer
describe how you would study turbidity or absorbance using a spectrophotometer
A
filter filters light of specific wavelength for example, at 540 nm and the spectrophotometer
measures the amount of light of this specific wavelength that passes through the sample
containing the cells. The higher the number, the cloudier the media indi
advantage of studying turbidity or absorbance using a spectrophotometer
quick and reliable
disadvantage of studying turbidity or absorbance using a spectrophotometer
it also counts both live and dead cells.
A flask whose content is transparent and is left overnight will turn cloudy because of
Microbial growth
Researchers describe the growth of bacteria, viruses and parasites as an
exponential growth
why do Researchers describe the growth of bacteria, viruses and parasites as an exponential growth?
because one cell (20, first generation) will divide to two cells (21, second generation), then
the two cells will divide to four cells (22, third generation) and so on
In the mathematical expression 2^n n represents
the generation number
Growth Curves can be plotted by using
a
semilogrithmic scale
When using a logrithmic scale, the slope
of the line represents the
rate of growth
is the time it takes for a population to double.
generation time
g =
time/number of generations For example, if it takes 60 minutes/6 generations, then it takes 10 minutes/generation.
For cells undergoing binary fission, the final cell number can be calculated if the
original cell
number and number of generations are known.
What is the total # cells present after 2 hours (120 minutes)?
Use final cell number = original cell number x 2^n = 400 cells/ml x 2^3 = 3200 cells/ml
If bacteria were removed from batch culture where there was a finite amount of media and
placed in new media, the first phase the bacteria would experience is the
lag phase
in what stage of the growth curve :bacteria are synthesizing components that they need for cell division. This phase is often
as a result of starvation, damage or step-down experiments (removing bacteria from rich media
and putting in nutrient-deprived me
lag phase
In the Exponential (Log) phase, the cells are the healthiest and are growing at
maximum growth rate possible
what stage of the growth curves is explained by , the rate of death of the cells is equal to the rate of the formation of
new cells so the population is steady
stationary phase
Bacterial cells enter stationary phase as they
run out of nutrients and accumulate waste
what stage of the growth curve can be explained by e there are more cells dying than are being formed and this varies in time-frame
death phase
In a _____, all cells in a population achieve a steady state. Fresh media
(nutrients) are supplied to the cells while spent media, wastes and excess microbes are
removed from a culture flask
continuous culture
Continuous culture allows
detailed study of bacterial physiology
since the researcher knows exactly how much media is being provided to the cells and how
much wastes are produced.
The relationship among the nutrient concentration in the vessel, generation time and bacterial
mass can be represented
graphically
As the nutrient concentration
increases, the bacterial mass also increases and the generation time
decreases
As the nutrient concentration
increases, the bacterial mass also increases however, the generation time decreases. Why?
This is
because the bacterial cells reproduce faster as they are better fed. Once washout is reached,
the bacteria are washed away.
Bacteria in the real world are in a mixed culture and live in ____known as a "city of
microbes".
biofilms
describe the steps in forming a biofilm
The first step in forming a biofilm is the attachment of the monolayer. Then
microcolonies are formed followed by the production of the exopolysaccharide (EPS). EPS is
the "glue" that hold the cells together and to the surface. A mature biofilm is then fo
Under stress, some bacterial species form
endospores
Clostridium and Bacillus
species are soil microbes that can produce dormant spores that are
heat-resistant, radiation
resistant as well as salt resistant.
starvation in bacteria cells initiates
an elaborate 8-hour differentiation process
where the cells go from a vegetative state (where they divide) to sporulation (genetic "trigger").
In sporulation, the cells are engulfed, spore coat is formed and dipicolinic acid synthesized.
Sporulation can be divided into discrete stages based primarily on
morphological appearance
Endospores have several layers to protect
the center that contains the DNA
Endospores resist environmental stresses such as
extreme temperature, drying, starvation, and
radiation.
Ancient halophile (salt-loving) endsopores have germinated from
crystals
Ancient halophile (salt-loving) endsopores have germinated from crystals .They have _____ present and a high content of ______which dries up the cells and
protects the DNA.
They have
dipicolinic acid (DPA) present and a high content of calcium
Endospores have less water than
vegetative cells
how to Endospores and Vegetative Cells differ?
high enzymatic activity and metabolism (oxygen uptake) which are indications
of life in vegetative cells while low enzymatic activity and low or absent metabolism indicates
dormancy in endospores
They also differ in their level of resistance. Resistance t