Paper

Praktikum Mikrobiologi

Microbial Growth
oleh:

Yuliana Sy
(0904103010068)
Kelompok B-2

JURUSAN TEKNIK KIMIA

FAKULTAS TEKNIK UNIVERSITAS SYIAH KUALA
DARUSSALAM, BANDA ACEH
2010

Growth is an orderly increase in the quantity of cellular constituents. It

depends upon the ability of the cell to form new protoplasm from nutrients
available in the environment. Microbial growth may be described as occurring in
different ways under different circumstances. In most bacteria, growth involves
increase in cell mass and number of ribosomes, duplication of the bacterial
chromosome, synthesis of new cell wall and plasma membrane, partitioning of the
two chromosomes, septum formation, and cell division. This asexual process of
reproduction is called binary fission. For unicellular organisms such as the
bacteria, growth can be measured in terms of two different parameters: changes in
cell mass and changes in cell numbers.
Methods for Measurement of Cell Mass
Methods for measurement of the cell mass involve both direct and indirect
techniques.
1. Direct physical measurement of dry weight, wet weight, or volume of cells
after centrifugation.
2. Direct chemical measurement of some chemical component of the cells such
as total N, total protein, or total DNA content.
3. Indirect measurement of chemical activity such as rate of O2 production or
consumption, CO2 production or consumption, etc.
4. Turbidity measurements employ a variety of instruments to determine the
amount of light scattered by a suspension of cells. Particulate objects such as
bacteria scatter light in proportion to their numbers. The turbidity or optical
density of a suspension of cells is directly related to cell mass or cell number,
after construction and calibration of a standard curve. The method is simple
and nondestructive, but the sensitivity is limited to about 107 cells per ml for
most bacteria.
(Kenneth Todar, 2008).
Because most bacteria grow by binary fission, doubling in cell number
usually occurs at the same rate that individual cells grow and divide. Binary
fission is the process by which most procaryotes replicate. Binary fission generally

involves the separation of a single cell into two more or less identical daughter
cells, each containing, among other things, at least one copy of the parental DNA.

Picture 1: Ilustration of Binary Fission

(Black, J.G., 1996).
There are three type bisection of cell.
1. Amitosis: cell reproduction where cell split directly without passing phases
bisection of cell. Bisectionof this cells having the character of prokariotik, for
example bacterium, blue algae
2. Mitosis: Cell reproduction where cell split pass regular steps, generally
happened at cells having the character of eukariotik. every mains cell in
mitosis which is diploid ( 2n) yielding two cell which is each remain to diploid
and measure up to clan which is equal to its mains cell.

Picture 2: Scheme bisection of cell by mitosis.

3. Meiosis: cell reproduction pass bisection phases like as mitosis, but in course of him
happened reduction chromosome.

Picture 3: Scheme bisection of cell by meiosis I

Picture 4: Scheme Bisection of cell by meiosis II

(Asnani, 2009).

Bacteria are all around us. Given good growing conditions, a bacterium
grows slightly in size or length, a new cell wall grows through the center forming
two daughter cells, each with the same genetic material as the parent cell. If the
environment is optimum, the two daughter cells may divide into four in 20
minutes (Anonimous, 2010).

Picture 5 Structure of Bacteria

Internal Structure: bacteria have a very simple internal structure, and no

membrane-bound organelles.

Nucleolid DNA in the bacterial cell is generally confined to this central

region. Though it isn't bounded by a membrane, it is visibly distinct (by
transmission microscopy) from the rest of the cell interior.

Ribosomes Ribosomes give the cytoplasm of bacteria a granular

appearance in electron micrographs. Though smaller than the ribosomes in
eukaryotic cells, these inclusions have a similar function in translating the
genetic message in messenger RNA into the production of peptide sequences
(proteins).

Storage Granules (not shown) Nutrients and reserves may be stored in the
cytoplasm in the form of glycogen, lipids, polyphosphate, or in some cases,
sulfur or nitrogen.

Endospore (not shown) Some bacteria, like Clostridium botulinum, form

spores that are highly resistant to drought, high temperature and other
environmental hazards. Once the hazard is removed, the spore germinates to
create a new population.

Surface Structure: Beginning from the outermost structure and moving

inward, bacteria have some or all of the following structures:

Capsule This layer of polysaccharide (sometimes proteins) protects the

bacterial cell and is often associated with pathogenic bacteria because it serves
as a barrier against phagocytosis by white blood cells.

Outer Membrane (not shown) This lipid bilayer is found in Gram negative
bacteria and is the source of lipopolysaccharide (LPS) in these bacteria. LPS is
toxic and turns on the immune system of , but not in Gram positive bacteria.

Cell Wall Composed of peptidoglycan (polysaccharides + protein), the cell

wall maintains the overall shape of a bacterial cell. The three primary shapes
in bacteria are coccus (spherical), bacillus (rod-shaped) and spirillum (spiral).
Mycoplasma are bacteria that have no cell wall and therefore have no definite
shape.

Periplasmic Space (not shown) This cellular compartment is found only in

those bacteria that have both an outer membrane and plasma membrane (e.g.
Gram negative bacteria). In the space are enzymes and other proteins that help
digest and move nutrients into the cell.

Plasma Membrane This is a lipid bilayer much like the cytoplasmic

(plasma) membrane of other cells. There are numerous proteins moving within
or upon this layer that are primarily responsible for transport of ions, nutrients
and waste across the membrane.

Appendages: Bacteria may have the following appendages:

Pili These hollow, hairlike structures made of protein allow bacteria to

attach to other cells. A specialized pilus, the sex pilus, allows the transfer of
plasmid DNA from one bacterial cell to another. Pili are also called fimbriae.

Flagella The purpose of flagella is motility. Flagella are long appendages

which rotate by means of a "motor" located just under the cytoplasmic
membrane. Bacteria may have one, a few, or many flagella in different
positions on the cell.
(Cellsalive, 2010).
Microbial growth curve was observed when microorganisms are cultivated

in batch culture. The culture was incubated in a closed vessel with a single batch
of medium. The curve was usually plotted as logarithm of cell number versus time
and usually has four distinct phases. Growth phase of mikroorganisme can be
differentiated to become 4 : Lag Phase, Exponential, stationary, and Death phase
(Anonimous, 2010).

Picture 6: Bacterial growth curve

The growth curve is a graphic representation of closed population of
bacteria overtime.
1. Lag Phase
At this stage, the cell synthesizing new components such as to replenish spent
materials or to adapt to new medium or other conditions. The length of the lag
phase is determined in part by characteristics of the bacterial species and in
part by conditions in the media - both the medium from which the organisms
are taken and the one to which they are transferred. Some species adapt to the
new medium in an hour or two; others take several days. Organisms from old
cultures, adapted to limited nutrients and large accumulated wastes, take
longer to adjust to a new medium than do those transferred from a relatively
fresh, nutrient-rich medium.
2. Log phase

Acceleration Phase (once after the cells have adapted to the new
environment; cell division occurs at increasing frequency until the
maximum growth rate reached)

Lag phase is followed by log phase during which binary fission occurs.
This phase of growth is called logarithmic or exponential because the rate
of increase in cell number is a multiplicative function of cell number. This
can be seen in a graph of cell number versus time where cell numbers

increase at ever increasing rates with time or generation; that is, the rate of
increase is a function of absolute cell number such that the more cells
present, the faster the population of cells increases in size (at least, during
log phase). During log phase, cells exhibit balanced growth, where cellular
constituents manufactured at constant rates relative to each other.
However, under certain condition (change in nutrient level and
environmental condition), unbalanced growth happens (rates of synthesis
of cell components vary relative to each other).

Deceleration Phase (when level of substrate decreases, it eventually

become limiting and no longer sustain maximum growth rate.

3. Stationary Phase
During this phase, total number of viable cells remains constant; mainly
because of metabolically active cell stop dividing or reproductive rate
balanced by death rate. Possible reason for entry into stationary phase
-

nutrient limitation

limited oxygen availability

toxic waste accumulation

critical population density reached

Starvation responses

morphological changes (such as endospore formation)

decrease in size

protoplast shrinkage

nucleoid condensation

production of starvation proteins

long-term survival

increased virulence

4. Death Phase
Death phase is a physiological point at which cell deaths exceed cell births.
More specifically, viable count declines. During the decline phase, many cells
undergo involution - that is, they assume a variety of unusual shapes, which
makes them difficult to identify.

Number of cells in each phase was estimated by using spectrophotometer.

In a spectrophotometer, light is transmitted through a dilution of the culture
(usually a 1:10 dilution). As microbe numbers increase, light passing through the
culture decreases. The output is often absorbance or optical density. Absorbance is
a logarithmic expression of the amount of light that gets through the culture. The
number of cells per absorbance unit is a known constant for most microorganisms
(Michael J. Waites, 2001).
Factors that Influence Bacterial Growth:
1. Physical requirements
a. Temperature
Bacteria have a minimum, optimum, and maximum temperature for growth
and can be divided into 3 groups based on their optimum growth temperature:
-

Psychrophiles

are

cold-loving

bacteria.

Their

optimum

growth

temperature is between -5C and 15C. They are usually found in the
Arctic and Antarctic regions and in streams fed by glaciers.
-

Mesophiles are bacteria that grow best at moderate temperatures. Their

optimum growth temperature is between 25C and 45C. Most bacteria are
mesophilic and include common soil bacteria and bacteria that live in and
on the body.

Thermophiles

are

heat-loving

bacteria.

Their

optimum

growth

temperature is between 45C and 70C and are comonly found in hot
springs and in compost heaps.
-

Hyperthermophiles are bacteria that grow at very high temperatures.

Their optimum growth temperature is between 70C and 110C. They are
usually members of the Archae and are found growing near hydrothermal
vents at great depths in the ocean.

Picture 7: Effect of temperature on bacterial growth

(Stephen Abendon, 1998).
b. Oxygen Requirements
Microorganisms show a great deal of variation in their requirements for
gaseous oxygen. Most can be placed in one of the following groups:
-

Obligate aerobes are organisms that grow only in the presence of oxygen.
They obtain their energy through aerobic respiration.

Example

Mycobacterium sp.
-

Microaerophiles are organisms that require a low concentration of oxygen

(2% to 10%) for growth, but higher concentrations are inhibitory. They
obtain their energy through aerobic respiration.

Obligate anaerobes are organisms that grow only in the absense of

oxygen and, in fact, are often inhibited or killed by its presense. They
obtain their energy through anaerobic respiration or fermentation. Example
Clostridium sp.

Aerotolerant anaerobes, like obligate anaerobes, cannot use oxygen to

transform energy but can grow in its presence. They obtain energy only by
fermentation and are known as obligate fermenters.

Facultative anaerobes are organisms that grow with or without oxygen,

but generally better with oxygen. They obtain their energy through aerobic
respiration if oxygen is present, but use fermentation or anaerobic
respiration if it is absent. Most bacteria are facultative anaerobes. Example
Saccharomyces sp.

Picture 8: Effect of oxygen on bacterial growth

c. pH
Most spoilage bacteria grow best near neutral pH. Pathogenic bacteria even
more narrow in tolerance range of near neutral. Yeast and moulds have much
greater tolerance to acidic (lower) pH. The optimum pH range is usually quite
narrow so that small changes in the pH can have large effects on the growth
rate of the organism. Effects of acids on organisms:

energy required to maintain cell's internal pH

enzyme activity affected

proteins, DNA, other molecules denatured

longer lag, less rapid growth

d. Osmosis is the diffusion of water across a membrane from an area of higher

water concentration (lower solute concentration) to lower water concentration
(higher solute concentration). Osmosis is powered by the potential energy of a
concentration gradient and does not require the expenditure of metabolic
energy. While water molecules are small enough to pass between the
phospholipids in the cytoplasmic membrane, their transport can be enhanced
by water transporting transport proteins known as aquaporins. The aquaporins
form channels that span the cytoplasmic membrane and transport water in and
out of the cytoplasm (see channel proteins below). A cell can find itself in one
of three environments: isotonic, hypertonic, or hypotonic . (The prefixes iso,
hyper-, and hypo- refer to the solute concentration).
(Anonimous, 2010).

2. Nutritional requirements
In addition to a proper physical environment, microorganisms also depend on
an available source of chemical nutrients. Microorganisms are often grouped
according to their energy source and their source of carbon
a. Energy Source
-

Phototrophs use radiant energy (light) as their primary energy source.

Chemotrophs use the oxidation and reduction of chemical compounds as

their primary energy source.

b. Carbon source
Carbon is the structural backbone of the organic compounds that make up
a living cell. Based on their source of carbon bacteria can be classified as
autotrophs or heterotrophs.
1. Autotrophs: require only carbon dioxide as a carbon source. An
autotroph can synthesize organic molecules from inorganic nutrients.
2. Heterotrophs: require organic forms of carbon. A Heterotroph cannot
synthesize organic molecules from inorganic nutrients. Combining their

nutritional patterns, all organisms in nature can be placed into one of four
separate groups: photoautotrophs, photoheterotrophs, chemoautotrophs,
and chemoheterotrophs.
3. Photoautotrophs use light as an energy source and carbon dioxide as
their main carbon source. They include photosynthetic bacteria (green
sulfur bacteria, purple sulfur bacteria, and cyanobacteria), algae, and green
plants. Photoautotrophs transform carbon dioxide and water into
carbohydrates and oxygen gas through photosynthesis.
4. Photoheterotrophs use light as an energy source but cannot convert
carbon dioxide into energy. Instead they use organic compounds as a
carbon source. They include the green nonsulfur bacteria and the purple
nonsulfur bacteria.
5. Chemolithoautotrophs use inorganic compounds such as hydrogen
sulfide, sulfur, ammonia, nitrites, hydrogen gas, or iron as an energy
source and carbon dioxide as their main carbon source.
6. Chemooganoheterotrophs use organic compounds as both an energy
source and a carbon source. Saprophytes live on dead organic matter while
parasites get their nutrients from a living host. Most bacteria, and all
protozoans, fungi, and animals are chemoorganoheterotrophs.
(Denikrisna, 2010).
c. Nitrogen source
Nitrogen is needed for the synthesis of such molecules as amino acids,
DNA, RNA and ATP. Depending on the organism, nitrogen, nitrates,
ammonia, or organic nitrogen compounds may be used as a nitrogen
source.
d. Minerals
1. Sulfur is needed to synthesisize sulfur-containing amino acids and
certain vitamins. Depending on the organism, sulfates, hydrogen sulfide,
or sulfur-containing amino acids may be used as a sulfur source.
2. Phosphorus is needed to synthesize phospholipids, DNA, RNA, and
ATP. Phosphate ions are the primary source of phosphorus.

3. potassium, magnesium, and calcium. These are required for certain

enzymes to function as well as additional functions.
4. iron is a part of certain enzymes.
5. trace elements are elements required in very minute amounts, and like
potassium, magnesium, calcium, and iron, they usually function as
cofactors in enzyme reactions. They include sodium, zinc, copper,
molybdenum, manganese, and cobalt ions. Cofactors usually function as
electron donors or electron acceptors during enzyme reactions.
e. Water
- Requirement for all living cells (70-90% water)
- Bacterial endospores and protozoa cysts can survive in low moisture
f. Growth factors
Growth factors are organic compounds such as amino acids, purines ,
pyrimidines, and vitamins that a cell must have for growth but cannot
synthesize itself. Organisms having complex nutritional requirements and
needing many growth factors are said to be fastidious.
(Tortora, 1995).

REFERENSI

Abendon, Stephen, Bacterial Gwrowth and Microbial Metabolism, diakses dari

http://mansfield.osu.edu/~sabedon/black06.htm, tanggal 12 Januari 2011,
pukul 17:52 WIB.
Anonimous, 2010, Bacteria Divide and Multiply, diakses dari http://www.cellsalive.com/ecoli.htm, tanggal 10 Januari 2011, pukul 16:29 WIB.
Anonimous, 2010, Microbial Growth Phase, diakses dari http://www.scribd.com/
doc/ 8775839/Microbial-Growth-Phase, tanggal 10 Januari 2011, pukul
16:33 WIB.
Anonymous, 2010, Factors affecting microbial growth, diakses dari http://www.
public.asu.edu/ ~pbaluch/courses/bio162/microbial_growth.pdf, tanggal 12
Januaei 2011, pukul 16:47 WIB.
Asnani, 2009, Pembelahan Sel, diakses dari http//asnani-biology.blogspot.com,
tanggal 10 Januari 2011, Pukul 17:51 WIB
Black, J.G., 1996, Microbiology, Principles and Applications, Third Edition,
Prentice Hall, Upper Saddle River, New Jersey.
Cellsalive, 2010, Bacterial Cell Structure, diakses dari http://www.cellsalive.com/cells/bactcell.htm, tanggal 10 Januari 2011, pukul 17:03 WIB.
Denikrisna, 2010, Pharmaceutical Microbiology Microbial Growth, diakses dari
http://denikrisna.wordpress.com/2010/11/01/pharmaceutical-microbiology
-%e2%80%93microbial%c2%a0growth/, tanggal 10 Januari
2011, pukul 17:49 WIB
Michael J. Waites, N.L. Morgan et all 2001, Industrial Microbiology: An
Introduction. Blackwell Science, London, UK
Todar, Kenneth, 2008, The Growth of Bacterial Populations, diakses dari http://
www.textbookofbacteriology.net/growth.html, tanggal 10 Januari 2011,
pukul 17:00 WIB.
Tortora, G.J., Funke, B.R., Case, C.L. 1995, Microbiology, An Introduction, Fifth
Edition The Benjamin/Cummings Publishing, Co., Inc., Redwood City.