Chapter 2

Structure, function, &multiplication of microorganism

-Based on the cellular complexity, which is revealed by electron microscope, microorganisms can be
classified as:
 Prokaryotes &
 Eukaryotes
Prokaryotic organisms
Prokaryote is a Greek word derived from two words: Pro-meaning primitive and Karyon meaning Nut shell
or nucleus .Generally, prokaryotic cells are primitive in nature and they are cells of lower life forms. The
nuclear material (DNA) is not enclosed with in a membrane but rather is distributed in masses through out
the cytoplasm. E.g. Mycoplasma, Bacteria, Rickettsiae
Eukaryotic organisms
Eukaryote is derived from two Greek words Eu (meaning true or real) & Karyon (meaning nucleus or nut
shell). The genetic material of Eukaryotic cells is surrounded by nuclear membrane forming a definite
nucleus e.g. Protozoa, fungi, Human. etc
Roll No Characteristic Feature Prokaryotic cells Eukaryotic cells
1 Nuclear membrane Not present Present
2 Chromosome number in the nucleus One in mass Greater than one
3 Mitochondria Absent Present
4 Division of cells By binary fission By Mitosis and Meiosis
5 Endoplasmic Reticulum Absent Present
6 Golgi apparatus Absent Present
7 Chloroplast Absent Present
8 Endocytosis & Exocytosis Absent Present

Bacteria
General property
 Typical prokaryotic cell
 Contain both DNA and RNA
 Most grow in artificial media
 Replication is by binary fission
 Contain rigid cell wall
 Sensitive to antimicrobial agent
Structure of Bacteria

Bacterial structure is considered at three levels.

1. Cell envelope
 Cell wall and cell membrane
2. Cellular elements enclosed with in the cell envelope:

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 Mesosome,ribosome, nuclear apparatus, and cytoplasmic granules.
3. Cellular element external to the cell envelope:
 Flagellum, Pilus and Glycocalyx(capsule)
.
1. Cell Envelope
A. Cell wall
Functions of Cell Wall
1. Provides shape to the bacterium
2. Gives rigidity to the organism
3. Protects from environment
4. Provides staining characteristics to the bacterium
5. Contains receptor sites for phage
6. Contains components toxic to host

B. Cell Membrane
o Also named cytoplasmic membrane & is a delicate
o It accounts for 30% of the dry weight of bacterial cell
o It is composed of 60% protein, 20-30% lipids and 10-20% carbohydrate.
Function of Cell Membrane
1. Regulates the transport of nutrients and waste products into and out of the cell.
2. Synthesis of wall components
3. Assists DNA replication
4. Secrets proteins
5. Caries on respiration
2. Cellular Element Enclosed with in the Cell Envelope
A. Mesosomes
 It is involved in DNA segregation during cell division and respiratory enzyme activity.
B. Ribosomes
 Cytoplamic particles which are the sites of protein synthesis
 It is composed of RNA (70%) and proteins (30%) and constitutes 90% of the RNA and 40% of the
total protein.
C. Cytoplasmic Granules
 Represent accumulated food reserves: glycogen etc.
D. Nuclear Apparatus.
 Bacterial genome consists of single molecule of double stranded DNA arranged in a circular form.
3. Cellular Element External to the Cell Envelope
A. Glycocalyx (Capsule)
-Capsule is composed of polysaccharide.
Features of Capsule
-Hardest part of the bacterium which is not necessary for viability but important in protection of the
bacterium from phagocytic cells.

B. Flagellum
-It is the organ of locomotion in bacterial cell and consists of filament on the surface of bacterial cell.
C. Pilli (Fimbriae)
-It is hair like structure located on bacterial surface.
 There are two types of pilli
 Common pili: The structure for adherence to cell surface
 Sex pilli: The structure for transfer of genetic material from the donor to the recipient
during the process of conjugation.

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 1. Classification of Bacteria
Bacteria can be classified on the basis of

1. Morphology (shape)

2. Staining (gram and other staining reaction)

3. Growths requirements

1. Morphology
Morphological classifications of bacteria when visualized under light microscope are:
A. Cocci (coccus) –Are bacteria that are round or oval in shape. They are found in single, pairs, chain or
cluster.
1.1 Monococcus: - Cocci occurring a singly.
1.2 Diplococci: - arranged in groups of two
1.3 Streptococci: - chain forming cocci
1.4 Staphylococci: - bunches of cocci or irregular groups or grapes of cocci
B. Rod shaped (Bacilli):- stick- like bacteria with rounded, tapered square or swollen ends.
C. Spiral shaped bacteria
Twisted or spiral shaped bacteria.
Borrelia, Treponema Leptospira
2. Staining
Staining is the process of coloring colorless objects or cells using stains (dyes)
Type of staining methods
1-Simple staining method
It is a type of staining method in which only a single dye is used.
-stain all types of bacteria and is used to observe the presence and shape of bacteria but not to differentiate.

2-Differential staining method

-used a number of reagent applied sequentially

- differentiate bacteria of different group based on the ability to acquire different color .e.g. gram stain
&acid fast stain.

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A. Gram’s stain

Developed by a Danish scientist, Christian Gram, in 1884.When applied it divides bacteria in to Gram
positive and Gram negative varieties.

 Procedures for Gram’s staining

Reagents Required:

 Crystal violet solution

 Iodine solution
 Alcohol (95 % ethanol)
 Safranin (counter stain)

Procedure

1. Make a thin smear on a slide and dry it in air

2. Cover the dry fixed smear with crystal violet solution and allow acting for 1 minute.
3. Remove excess stain briefly rising the slide with gently flowing tap water.
4. Flood the smear with iodine solution and allow standing for 1min.
5. Rinse off the iodine with tap water & apply a decolizer solution just until no more color is being
washed from the smear.
6. Shake off any excessive water and apply safranin counter stain solution for one min.
7. Wash the slide with tap water, blot dry without rubbing.
8. Examine under the microscope.

B. Acid fast stain (Ziehl-Neelson)

Mycobacteria, the mycoplamas,the rickettsias and some of the spiral bacteria do not stain by grams method
and for them other stains or techniques must be used.The mycobacteria are made visible by a method called
ziehl-Neelson.When applied,it divides bacterias in to Acid fast and non-acid fast.

Since acid fast bacilli's possess high amount of waxy substances in their cell wall, they retain the color of
basic fuchsin (reddish in color).

Procedure

Reagents used in acid fast staining.

o Carbolfuschsin
o Acid alcohol
o Methelene blue or malachite green

Staining

 Prepare smear from clinical sample and air dry. Then heat fixation.
 Cover the smear with carbolfuschsin and use heat until steam rise. Then wash.
 Cover with 20 % sulfuric acid / 3% HCl in 95 % Ethanol (acid alcohol) then wash.
 Cover the smear with methylene blue/ malachite green for one minute. Then wash with distilled water.
 Dry the smear and observe under microscope.

E.g. Acid fast bacilli: Mycobacterium tuberculosis, Mycobacterium leprae

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iii. Classification of bacteria based on their growth requirement

Depending on growth requirement, bacteria's can be classified in to different groups based on the
following:

1. Air requirement

Oxygen requirement: On the presence of O2 bacteria can be classified in to:

 Obligatory (strict) Aerobes – bacteria which needs free oxygen for their growth.
 Obligate or strict anaerobes- bacteria which only grow in absence of free O 2. They have oxygen
sensitive enzymes.
 Facultative anaerobes – these are organisms which can grow either in the presence or absence of O2
 Microaerophilic – are aerobic bacteria's which need small amount of O2 for growth.
 Carboxyphilic (capnophylic) bacteria's – require an atmosphere composed of oxygen and raised level
of CO2

2. PH requirement.

pH is the measure of acidity or alkalinity of a solution or a medium.

o PH below 7 is acidic
o PH above 7 is alkaline (basic)
o PH=7 is neutral

 Most bacteria's reproduce at a pH range of 6 -8.

 Few bacteria grow at a pH higher than 8.0 and they are called alkalophilic. E.g. Vibrio cholerae.
 Those bacteria's which can grow at a pH less than 7.0 are called acidophilic. E.g Lactobacillus.
 There are some bacteria's which grow at a pH between 4.0 and 9.0 which are called neutrophilic.

3. Temperature

There is wide range of temperature within which all bacteria's can grow. This range can influence
enzymatic activity.

o Mesophilic – those bacteria's which can grow at a temperature between 20 oC - 50 oC.

o Psychrophilic – those bacteria's which can grow at a temperature between 0oC – 20 oC.
o Thermophilic – those bacteria's which grow between 50 oC – 100 oC.
 Most pathogenic bacteria's are mesophils.

4. Moisture

Most bacteria need optimum amount of moisture but there are some bacteria which need high amount of
moisture.

E.g. T. palladium, N. gonorrhea

5. Nutritional requirement – There are different types of nutritional requirement of bacteria's depending
on the nature of bacteria's.

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 Depending on their nutritional requirement bacteria can be classified

 Autotrophes (simple)- Free-living, non-parasitic bacteria which use carbohydrate as carbon source the
energy needed for their metabolism can be obtained from sun light and inorganic compound
 Heterotrophs (complex)– In addition to the simple nutritional requirement, parasitic bacteria require
more complex organic compounds as their source of carbon and energy

N.B. Human phatogenic organisms are heterotrophs

 Halophilic – Those organisms that live (grow) on high salt concentration.

E.g. Staphylococcus aurous needs 7.5-g/l salt concentrations. Normal requirement is 5 g/l salt

Bacterial Genetics

 Genetics is a study about inherited characters.

 Bacterial inherited characteristics are encoded in a single molecule of DNA arranged as a double
enclosed loop.
 Bacterial genetic material is organized as:

1. Chromosome
2. Extra chromosome (Plasmid)

1. Chromosome: bacterial chromosome is circular double stranded DNA attached to bacterial cell
membrane
2. Plasmids: are self-replicative extra chromosomal DNA molecules.

Plasmids are not essential to the life of the cell but they may have selective advantage for these
organisms like resistance to antibiotic.

Genetic variation in Bacteria

Genetic Variation in Bacteria can occur by

1. Mutation
2. Gene-Transfer

1. Mutation

Bacterial mutation occurs when the information in a bacterial chromosome is altered by different means
like chemical or radiation.

There are two types of mutation:

1. Spontaneous mutation
2. Induced mutation

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 Spontaneous Mutation

- Spontaneous mutation are those that takes place in nature with out human intervention or
other identifiable causes

E.g. Penicillin- resistant strains of Neisseria gonorrhea may have emerged due to spontaneous
mutation

Induced mutation

- Are those mutations in which the cause can be identified

 Exposure to chemical or physical agent

 From treatment with ultraviolet light

1. Gene-Transfer

There are three types of gene transfer that alter the DNA content of bacteria

1. Transformation
2. Transduction
3. Conjugation

A. Transformation

It occur when fragment of exogenous bacteria DNA are taken up and absorbed into recipient
bacterial cells

This can result in:

 Change in pathogencity of the bacterium

 Change in antibiotic sensitivity pattern of the bacterium

B. Transudation

It occurs when fragments of chromosomal DNA is transferred or transduced from one bacteria to
another through phage particle (phages are viruses which infect bacteria).

C. Conjugation

Conjugation occurs when plasmid DNA is transferred form donor bacterium to recipient
bacterium by direct contact via sex pillus.

2.3. Viruses

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Viruses are organized association of nucleic acid with a protective shell of protein units
(envelop or capsid). Viruses are very small in size ranging 20 to 300 nm in diameter. Unlike
most bacteria, fungi and parasite, viruses are obligate interacellular parasites that depend on the
biochemical machinery of the host cell for replication.

Viral structure

Viral structural parts include:

1. Nucleic acid core: The nucleic acid core consists of viral genome located internally and can
be either single or double stranded DNA or RNA molecule.
2. Capsid: it is a protein coat that surrounds the nucleic acid genome.
3. The envelope: Is the outer most coat of the virus that is used for attachment with host cell
receptor. Viruses may have envelope or can be naked capsids. Naked viruses are resistant
than enveloped viruses.

Properties of viruses

 Viruses are filterable agent

 Viruses are obligate intracellular parasites
 Viruses cannot make energy or proteins independent of a host cell
 Viral components are assembled and do not replicate by division

Classification of viruses

-Based on their nucleic acid, Viruses can be grouped into DNA or RNA Viruses.

-Based on their packaging structure, viruses can be grouped into capsid or enveloped viruses.

DNA virus RNA virus

Enveloped capsid Enveloped Capsid

E.g. hepatitis B e.g. papiloma virus e.g. retrovirus (HIV) e.g. Rota
virus

2.4. Rikettsiae & Chalamydiae

o The rickettsiae and Chlamydiae are obligate intracellular organisms.

o Chalamydiae are aerobic and the rickettsiae are anaerobic. Both are bacteria that are small,
pleomorphic coccobacillary.
 Chalamydia trachomatis: It causes trachoma; the disease is transmitted from infected
person by contaminated fingers, towels, or by houseflies.
 Rickettsia: Rickettsial disease is referred as arthropod transmitted disease. Thick, lice,
fleas, and mites are vectors of a variety of diseases caused by rickettsias.
2.5. Mycoplasamas

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 The smallest living micro-organism capable of independent existence
 Highly pleomorphic bacteria due to absence of rigid cell wall, instead bounded by a “unit
membranes”
 Completely resistant to penicillin and cephalosporin but sensitive to Tetracycline and
Erythromycin

2.6. Fungi
 Fungi are saprophytic or parasitic eukaryotic microorganisms distinct from plant and animals
 The diseases caused by fungi are called Mycoses.
Morphologic forms of Fungi
Yeasts
 Yeast are unicellular and reproduce by extrusion of buds from the mother cell
Molds
 Molds are filamentous branching stands of connected cells which form a hypha (a
collection of hyphae comprise a mycelium)
 Colonies of mycelia fungi have cottony or velvety surface which is dry and easily broken
a part
Dimorphic fungi
 Having yeast and filamentous shape
Spore formation
 Fungi reproduce asexually by budding forming spore, which are structures that are
capable of surviving under adverse environmental conditions