Transverse binary fission is the most
common asexual reproductive process whereby a single cell divides into two
after developing a transverse cell wall.
Species of the genus Streptomyces
produce many reproductive spores; each spore gives rise to a new organism.
Related bacteria (genus Nocardia) produce extensive filamentous growth which is
followed by fragmentation of the filaments into bacillary or coccoid cells.
Multiplication leads to an increase in
the number of individual cell making up a population or culture. Growth
denotes the increase in number beyond that present in the original inoculum.
Thus, if we start with a single bacterium, the increase in population is by geometric
progression.
Generation is the
doubling of cell in number. The time interval required for the cell to divide
is known as generation time. Not all bacteria have the same
generation time. The generation time is dependent upon the nutrients in the
medium and the physical conditions. Growth rate is expressed in terms of
generation per hour.
When a given number of cells are
inoculated into a fresh medium, the bacterial population is determined
intermittently during an incubation period of 24 hours (more or less) and the logarithms
of the cells versus time are a representation of population changes in the
growth of a culture.
Section of curve Phase Growth rate
A Lag Zero
Acceleration
Increasing
B Exponential Constant
Retardation Decreasing
C Maximum Zero
Stationary
D Decline Negative (death)
Four major phases of bacterial
growth curve
1. Lag
phase
– adaptation period
Enzymes and
intermediates are formed and accumulate until they are present in
concentrations that permit their growth to resume. The cells are metabolizing
but there is a lag in cell division.
2.
Logarithmic or
Exponential phase
The cells
divide steadily at a constant rate. Growth rate is maximal during this phase.
The population is most nearly uniform in terms of chemical composition of
cells, metabolic activity and other physiological characteristics.
3.
Maximum
stationary phase
There is a
stopping of growth completely attributed to the exhaustion of nutrients and
production of toxic products during growth. The population remains constant for
a time as a result of complete stopping of division or the balancing of
reproduction rate by an equivalent death rate.
4.
Death phase or
period of decline
After
stationary phase, bacteria may die faster than new ones produced, if some cells
are still reproducing. Depletion of nutrients and accumulation of inhibitory
products contributes to bacterial death. Bacteria die at different rates, just
as they grow at different rates.
An appreciation of the normal growth
curve is important; it must be understood that the cell are young and actively
metabolizing during some phases of growth while in other phases they are dying,
so there may be a great structural and physiological differences between cells
taken at different times. In general, cells in the logarithmic phase of growth
are commonly used for studies of metabolism because they are the most uniform
and in a more clearly defined condition than any other bacterial cells.
There are many aspects of research
relating to cell growth, organization and differentiation, with these aspects;
it is desirable to have an entire population of cells in the same stage of
growth cycle. The growth pattern in which all the cells will divide at the same
time is known as synchronous growth. One can synchronize a
population by manipulating the physical environment or the chemical composition
of the medium.
Steady state or balanced growth is a
condition wherein the bacterial population is maintained in the exponential or
log phase. A bacterial population in this condition is used both in
experimental research and in industrial processes. Devices for this are the turbidostat
and the chemostat. In both devices, fresh medium is
allowed to enter the culture vessel at the same rate as medium spent is removed
to keep the culture at a constant volume.
Growth can be measured
quantitatively by:
a. Cell
count – directly by microscopy or an electronic particle counter or indirectly
by a colony count to determine viable bacteria.
b. Cell
mass – directly by weighing or by measuring nitrogen content or indirectly by
turbidity.
c. Cell
activity – indirectly by relating the degree of biochemical activity to the
size of the population.
Summary of methods for
measuring bacterial growth
Method
Applications
Microscopic count Enumeration of
bacteria in milk and vaccine
Plate count Enumeration of bacteria in
milk, water, foods,
soil,
cultures
Membrane or molecular filter same as plate count
Nitrogen determination measurement of cell crop from
heavy culture
suspensions
to be used for research in metabolism
Weight determination same as for nitrogen
determination
Turbidimeteric measurement microbiological assays, estimation of
cell crop in
broth,
culture aqueous suspension
Measurement of biochemical microbiological assays
activity
ENZYMES AND
THEIR REGULATION
To live, grow and reproduce, a cell
must be capable of performing an immensely of chemical changes, and these
chemical changes lies in the activity of the enzymes. In a sense, the enzymes
may be regarded as the working part of the cell.
There are two types of enzymes
based on the site of action
1.
Intracellular
or endoenzymes
– functioning in the cell. It synthesizes cellular material and also performs
catabolic reactions which provide the energy requirements of the cell.
2.
Extracellular
or exoenzymes
– functioning outside the cell. It performs whatever changes are necessary on
the nutrients in the medium to allow the foods to enter the cell.
Based on the presence of
substrate and enzyme formation, bacterial enzymes are divided:
1.
Constitutive – these are
always produced by the cell, independently of the composition of the medium in
which it grows.
2.
Adaptive
(induced)
– these are produced by the cell only in response to the presence of a
particular substrate; they are produced only when needed.
The growing culture technique
is used routinely for the characterization of the enzymatic activities of
microorganisms. Results of such test provide information necessary for their
identification. The resting cell technique and cell free preparation are
principally used in research work, the purpose of which is to determine how the
organisms accomplishes each specific change.
Procedure for growing culture
technique:
1. Inoculation
of the bacteria into a medium containing the substrate.
2. Incubation
of the medium containing the bacteria for 1 or more days.
3. Examination
for a change or disappearance of the substrate and presence of end products.
The living cell neither synthesizes
nor catabolizes more material than is required for normal metabolism and growth,
all this cells for precise control mechanisms of cellular metabolism. The
control of cellular metabolism eventually involves the regulation of enzyme
activity. Generally speaking, enzymes can be controlled or regulated in two
ways:
1. Genetic
control –
inducing and suppressing enzyme synthesis is at the genetic level.
2. Direct
control of catalysis – by altering the concentrations of substrate or
reactants or by coupling with other processes which usually implies regulation
of the ligands which do not participate in the catalytic process itself.
Feedback
inhibition
– is one type of this control by which the end product of a series of metabolic
reactions inhibits the activity of an earlier enzyme of the sequence.
BACTERIAL
METABOLISM
The term metabolism denotes all the
organized chemical activities performed by a cell, which comprises two general
types: (1) energy production (2) energy utilization
Energy production
Most bacteria obtain energy by
carrying out chemical reactions which liberate energy. Some forms of life such
as green plants, can utilize radiant energy and are designated as phototrophs.
Others rely upon oxidation of chemical compounds and they are called chemotrophs.
The systems in bacteria that transform
chemical and radiant energy into a biologically useful form include:
1. Respiration – in which
the molecular oxygen is the ultimate electron acceptor. The pathway of aerobic
dissimilation is exceedingly complex. The most important respiratory mechanism
for final oxidation is the tricarboxylic acid cycle of Krebs, which together
with the known reactions of glycolysis, can account for the complete oxidation
of glucose.
Kreb cycle – an enzyme
system which converts pyruvic acid to CO2 in the presence of O2
accompanying release of energy which is trapped in the form of ATP (adenosine
triphosphate) molecules.
2. Fermentation – the
foodstuff molecule is usually broken down into two fragments, one which is then
oxidized by the center.
The major
route of glucose catabolism in most cells is glycolysis or the Embden–Meyerhoff
pathway, an anaerobic process of glucose dissemination, which may occur
by sequence of enzyme catalyzed reactions, to pyruvic acid.
Although the
basic pathway is the same for all types of cells, the properties of certain
enzymes are not uniform in all species or cell types. Such variations are
introduced for purpose of cellular distinction and control of specific steps in
the pathway.
Respiration in
the presence of oxygen, glucose is changed into CO2 and water. It is
much more efficient process than fermentation because many molecules of ATP are
generated. Fermentation in the absence of oxygen, glucose is changed into
different end products and only a few molecules of ATP is generated
3. Photosynthesis – a process
whereby microorganisms with chlorophyll obtain energy from the environment in
the form of light.
Energy utilization
Once energy is obtained, bacteria as
well as other organisms utilized it in various ways:
1. For
the biosynthesis of new cell components
2. For
the maintenance of physical and chemical integrity of the cell.
3. For
the activity of the locomotor organelles
4. For
transporting solutes across membranes
5. For
heat production
Death – is the
reversible loss of the ability of the cell to reproduce.
Empiric test
of death –
is determined by culturing the cells on solid media. A cell is considered dead
if it fails to give rise to a colony.
MICROBIAL
GENETICS
From the point of view of genetics, a
characteristic of all forms of life is the general uniformity or “likeness” in characteristics
of the progeny and parent. A particular organism, however, may not exhibit the
same characteristics when grown under a variety of different environmental
conditions. A deviation from the parent form in the bacteria of the same
species growing under the different or identical condition is known as variation.
It occurs as a result of:
1. Genotypic
changes
– involves alteration of genes
a.
Mutation
Any gene is
capable of changing or mutating to a different form, so that it causes an
altered characteristic. The act of mutating is known as mutation
or it is an alteration in form or qualities. In mutation, there is a permanent transmissible
change in characters of offspring from those of parents. It occurs in two ways:
(1)
Spontaneously – under
natural conditions due to tautomeric shift of electrons in purine or pyrimidine
base
(2)
Induced – due to
exposure of cells to some mutagenic agents such as base analogue, alkylating
agents, ultraviolet radiations, etc.
b.
Recombination
It occurs when
a portion of the genetic material from a donor is transferred to a recipient
cell. There are three processes by which recombination takes place:
(1)
Transformation – soluble DNA
from the donor cell is released into the surrounding medium and taken up by a
recipient cell.
(2)
Transudation – fragment of
the donor chromosome is carried to recipient cell by a temperate bacteriophage
(virus–eating bacteria)
(3)
Conjugation–plasmid
mediated
– two bacteria actually join together to permit transfer of the genetic
material through the pilus.
Plasmid – is a small,
extrachromosomal genetic unit which cannot be integrated into the chromosome
and not transferred by an infectious process (unless taken up by a generalized
transducing bacteriophage).
They are
clinically significant because:
(a) They
carry genes for resistance to therapeutic agents
(b) They
lead to emergence of strains to new combinations with antigenic and virulent
factors.
Their presence
is detectable only when the genes they carry confer new properties on the host.
2. Phenotypic
changes
– changes are imposed by the environment without genetic change. It is less
stable than genotypic change and is not inherited.
a.
Morphological
modifications
(1)
Cellular
variation
(a) Large
capsules are produced by certain species of bacteria when grown in milk and
none at all when grown in nutrient broth.
(b) Spores
are formed when certain species of bacteria are placed in an environment that
is unfavorable for their growth.
(c) Flagella
are removed from a certain bacteria when placed in a medium containing alcohol.
(2)
Colony
variation
Colony is a group of
bacteria formed from the reproduction of a single organism and generally
visible to the naked eye.
Dissociation is a kind of
bacterial variation in which there is a change in the observable
characteristics of growth in colonies resulting in the production of a new one.
A change from S (smooth) to R (rough) forms is an example of dissociation.
Other types are:
(a) M
(mucoid) – viscous and slimy
(b) H
(spreading) to O (nonspreading)
(c) D
(dward or diptheroid)
Pleomorphism is variation
in size, shape and appearance of bacteria of the same species growing under
favorable conditions.
Involutions or
degenerative forms – these are abnormal forms assumed by
microorganisms growing under unfavorable conditions.
b.
Physiological
variation
Adaptation – variations (changes
in bacterial make–up) that represent physiologic adjustment to the environment
(1) Ability
to decompose sugars
(2) Variation
in nutritive requirement
(3) Susceptibility
to infection by viruses
(4) Immunological
characteristics
(5) Virulence
Attenuation – an
important form of adaptation which indicates a loss in disease–producing
ability to a given organism.
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