MICROBIOLOGYIntroduction:

‘Microbiology’ should be an easy word to define: the science

(logy) of small (micro) life (bios), or to put it another way,

the study of living things so small that they cannot be seen

with the naked eye.

Microbiology is the science that deals with the study of

micro-organisms, their activates and their influences on

different aspects of life.

The following list of some of the general areas in which the

expertise of a microbiologist might be used:

1-Medical laboratory Sciences

2- Medicine

3- Environmental science

4- Food and drink production

5- Fundamental research

6- Agriculture

7- Pharmaceutical industry

8- Genetic engineering.

Microbiology comprises many disciplines, that are categorized

on either taxonomic or functional bases:

(1) Taxonomic disciplines of microbiology are:

Bacteriology (Study of bacteria)

Mycology (Study of fungi)

Virology (Study of viruses)

Parasitology ( Study of parasite) (now in separated section)

(2) Functional disciplines of microbiology are:

Immunology (host reaction to micro-organisms)

Medical Microbiology (role of organism in infectious

disease)

Genetic engineering

Historical background:

• WE NEED TO KNOW OUR PAST SO WE KNOW THE DIRECTION THAT

WE NEED TO FOLLOW IN THE FUTURE.

•Microorganisms had been on the Earth for some 4000 million years, when

Antoni van Leeuwenhoek started out on his pioneering microscope work in

1673. Leeuwenhoek was an amateur scientist who spent much of his spare

time grinding glass lenses to produce simple microscopes.

•the Italian Francesco Redi (1626–1697) showed that the larvae found on

putrefying meat arose from eggs deposited by flies, and not spontaneously as a

result of the decay process. This can be seen as the beginning of the end for

the spontaneous generation theory,

•Pasteur noticed that when lactic acid was produced in wine instead of alcohol,

rod-shaped bacteria were always present, as well as the expected yeast cells.

This led him to believe that while the yeast produced the alcohol, the bacteria

were responsible for the spoilage (swan-necked flasks).

•The first proof of the involvement of bacteria in disease and the definitive

proof of the germ theory of disease came from the German Robert Koch. In

1876 Koch showed the relationship between the cattle disease anthrax and

a bacillus which we now know as Bacillus anthracis.

•These results led Koch to formalise the criteria necessary to prove a causal

relationship between a specific disease condition and a particular

microorganism. These criteria became known as Koch’s postulates, and are

still in use today.

Koch’s postulates:

(1) The microorganism must be present in every instance of the disease

and absent from healthy individuals.

(2) The microorganism must be capable of being isolated and grown in pure

culture.

(3) When the microorganism is inoculated into a healthy host, the same

disease condition must result.

(4) The same microorganism must be re-isolated from the experimentally

infected host.

•Table (1) The discovery of some major human pathogens

•Year Disease Causative agent Discoverer•1876 Anthrax Bacillus anthracis Koch

•1879 Gonorrhoea Neisseria gonorrhoeae Neisser•1880 Typhoid fever Salmonella typhi Gaffky

•1880 Malaria Plasmodium sp Laveran•1882 Tuberculosis Mycobacterium tuberculosis Koch

•1883 Cholera Vibrio cholerae Koch•1883/4 Diphtheria Corynebacterium diphtheriae Klebs & Loeffler

•1885 Tetanus Clostridium tetani Nicoaier & Kitasato•Year Disease Causative agent Discoverer

•1886 Pneumonia (bacterial) Streptococcus pneumoniae Fraenkel•1892 Gas gangrene Clostridium perfringens Welch & Nuttall

•1894 Plague Yersinia pestis Kitasato & Yersin•1896 Botulism Clostridium botulinum Van Ermengem

•1898 Dysentery Shigella dysenteriae Shiga•1901 Yellow fever Flavivirus Reed

•1905 Syphilis Treponema pallidum Schaudinn & Hoffman•1906 Whooping cough Bordetella pertussis Bordet & Gengou

•1909 Rocky Mountain spotted fever Rickettsia rickettsii Ricketts

Leeuwenhoek’s microscope

The lens (a) was held between two brass plates and used to view the

specimen, which was placed on the mounting pin (b). Focusing was

achieved by means of two screws (c) and (d).

Pioneers in Microbiology: • Aristotle (384–322BC) believed that life arose from muck, decaying

food, warm rain, or even dirty shirts.• Antony van Leeuwenhoek (1632–1723)

used a primitive microscope to observe stagnant water, hay infusions, and scrapings from the teeth.

• John Tyndall (1820–1893) discovered that some bacteria existed in two forms: a heat stable form and a heat-sensitive form.

THE GOLDEN AGE OF MICROBIIOLOGY • Mid to late 1800’sHISTORY CONT.

• Louis Pasteur– France

– Fermentation—with the final result being pasteurization.• Heating wine and milk to 145.5 degrees to kill bacteria. Without killing

this bacteria, both the wine and the milk would sour. Louis Pasteur cont.– Developed immunizations for anthrax in cattle and rabies in humans.

• Robert Koch– Koch’s Postulates

• An organism must be present in ALL cases of the disease.• The organism must be isolated in pure culture.

• The organism can be removed from the pure culture and placed in another human or experimental animal and the same symptoms are

developed.• The organism can be removed from this new person and grown again in

pure culture.• His postulates prove that certain organism cause certain diseases.

• Ignaz Semmelweis—Vienna and OliverWendell Holmes—America– Discover the importance of hand washing prior to surgery and then between patients.

Other Introduction:

The word Microbiology is a Greek one: It is the study of very small

organisms (Microorganisms) that can not be seen by naked eye but by

Microscope.Micro = very small

bio = lifelogy = Science

These microorganisms include four major groups:

•Bacteria: studded under the name of Bacteriology.

•Fungi: studded under the name of Mycology.

•Viruses: studded under the name of Virology.

•Protozoa: studded under the term of Parasitology & Protozoology.

Why do we study Microbiology?

Because almost all diseases are caused by microorganisms.

There is a classification for these major groups of microorganisms which

can be distinguished on the bases of their structure and the complex of

their organization.

A- Prokaryotes: Which included Bacteria and blue green algae.

Characteristics of prokaryotes:

1- Very small in size.

2- Lack of nuclear membrane.

3- Lack of mitochondria.

4- Multiplication is by binary fission.

5- Has a single piece of genetic material known as plasmid.

B- Eukaryotes: Which included Fungi and Protozoa.

Characteristics of eukaryotes:

1- Large size.

2- Lack of plasmid.

3- Posses mitochondria.

4- Multiply by sexual multiplication & others (mitosis).

5- Posses nuclear membrane.

Classification of Micro-organisms:

In any discussion on biological classification, it is impossible to avoid

mentioning Linnaeus, the Swedish botanist who attempted to bring order to

the naming of living things by giving each type a Latin name. (Carl von Linn´e!

It was Linnaeus) who was responsible for introducing the binomial system of

nomenclature, by which each organism was assigned a genus and a

species.

The science of taxonomy involves not just naming organisms, but

grouping them with other organisms that share common properties.

In the 20th century, an increased focus on the cellular and molecular

similarities and dissimilarities between organisms led to proposals for further

refinements to the three-kingdom system. One of the most widely accepted

of these has been the fivekingdom system proposed by Robert Whittaker in

1969. This took into account the fundamental difference in cell structure

between procaryotes and eucaryotes, and so placed procaryotes (bacteria)

in their own kingdom, the Monera, separate from single-celled eucaryotes.

Another feature of Whittaker’s scheme was to assign the Fungi to their own

kingdom, largely on account of their distinctive mode of nutrition.

The agents of human infectious belong to five major groups of micro-

organisms ;bacteria, fungi, viruses protozoa and helminthes.

Based upon cell characteristic, living organism are recognized as having

Euocaryotic or procaryotic cell type.

Bacteria belong to the procaryotic, fungi and protozoa are Euocaryotic

cell type. Viruses are classified by some scientist as kingdom viruses they

are acellular (non- cellular)

 Euocaryotes procaryotes

Size Larger Relatively small

Structure of cell Complex Primitive

DNA content Nucleus Nucleoid

Nuclear membrane Present Absent

Mitochondria Present Absent

Multiplication Mitotic Binary fission

Enzyme system

Golgi bodies

Complex

Present

Simple

Absent

Respiration Via Mitochondria Cytoplasmic membrane

Differences between Euocaryotes and procaryotes

The kingdom procaryote is further divided into division classes

orders families genera species strain

(subspecies)

Practically, only the name of genus and species are used

the generic (genus) name is always given a capital letter

the specific (species) name is given a small letter

the generic and specific name are italicised, or, if this isn’t possible,

underlined.

Introduction to Bacteria

Bacteria form a large group of unicellular parasitic, saprophytic and free-living

microorganism, varying in size from .01 – 10 µm long.

They have simple cell structure (procaryotic), contain both DNA &RNA and multiply by

binary fission They are classified by their morphology, staining reaction, culture characteristic,

biochemical reaction, antigenic structure and genetic composition using specialized

molecular biology techniques.

Morphology of Microorganisms:Morphology of Bacteria:-

1- Cocci: round or spherical in shape; singular known as

Coccus. It's diameter ranges between 0.5 – 1 µm. When

microorganism occurs as single it is known as Micrococcus

monococcus = single cell.When it occurs in pairs, it is known as Diplococci e.g. Meningococci which cause meningitis.When it occurs in chain which can be short or long, it is known as Streptococci e.g. Streptococcus pyogenes which cause throat inflammation.

When it occurs in clusters it is known as Staphylococcuse.g. Staphylococcus aureus (golden colour).

These arrangements are determined by the orientation and degree of attachment of the bacteria at the time of cell

division.

2- Rods:in Latin are known as Bacilli. Rod = Bacillus ; it has a stick like shape

and the end may be round cut or square cut (Value of identification).

When rod is single, it is known as bacillus or may be chain known as

streptobacillus

Sometimes the rods may occur in chains but

branching, some have the shape of Chinese

letters and is known as club shaped e.g.

Corynebacterium diphtheriae which causes

diphtheria.

3-Vibrios: It has a comma or curved shape ،،It is known as Vibrio and e.g. Vibrio cholerae

which causes cholera.

4- Spirillum and Spirochetes:

have the shape of spiral e.g.Treponema palidum

which causes syphilis.

Molecular Structure of Bacteria

1- Cell Wall: is a component of bacterial cell which

is outside the cell, it is a rigid organelle and also it

gives the bacteria it's shape and also prevents

expansion of cell membrane (protects the cell against

osmotic pressure). The other name of the cell wall is

known as peptidoglycan which is composed of:

polysaccharides:

the main polysaccharides which are found

in the composition are:

N – acetyl glucose amine.

N – acetyl muramic acid.

and this peptidoglycan is only found in the cell wall

of bacteria. It is a target for antibiotics to act on.

Some antibiotics act on the cell wall e.g. penicillins

and cephalosporins. This is known as selective

toxicity.

Defect or obstruction of the cell wall will give:

1. Protoplast: is a G+ve bacteria with defective cell wall which is

killed by lysozymes or antibiotic (penicillin).

2. Spheroplast: Is a G-ve bacteria which lost its cell wall

completely or partially due to the action of lysozymes or antibiotic

(penicillin).

3. L-forms: It may be a G+ve or G-ve bacteria loss their cell wall due to

mutation. They are able to grow and divide or multiply and this is possible in a

hypertonic or concentrated media or isotonic in order to retain its contents and do

not release them. In this medium shrinking may happened but it does not affect.

But in hypotonic media it will expand and then explode.

2. Cell Membrane:

It is very important structure in the bacterial cell, also called

cytoplasmic membrane or sometimes protoplasmic

membrane. Chemically cytoplasmic membrane is composed

of bilayers of phospholipids.

Basic functions of cell membrane are :

1\ to control the movement of substances.

2\ to secrete hydrolytic enzymes.

3\ responsible for secretion of transport protein and proteins

involved in cell wall synthesis.

3- Cytoplasm: which contains:

a. Genetic Material: a single piece of DNA

without nuclear membrane.

b. Ribosomes: which are known as protein

making organelle.

c. Mesosomes: Which are attached to the

cell membrane and is thought that it is

associated in cell division (binary fission).

Specialized products outside the cell wall

1- Capsule:

this is a layer of loose slime material which

surrounds some bacterial cells. The capsules are

composed of mainly of polysaccharides or peptides.

They resist phagocytosis and so their presence on a

bacterium is associated with virulence. They are

identified by negative staining due to their low affinity

for simple staining.

* e.g. of capsulated bacteria: Klebsiella pneumoniae

(polysaccharide)Bacillus anthraces (poly – D – glutamic acid)

* The capsule is used for rapid and accurate diagnosis where agglutination reaction happens between the antibodies and the antigen (capsular antigen).

2- Flagella:

These are filaments that originate from the cytoplasm.

They function as organs of motility. They are therefore seen

only in organisms that are motile. They are made of protein

(Flagellin). They have characteristic patterns of arrangement

on the bacterial cell:

A) Monotrichate: one flagellum at one pole.

b)Amphitrichate: one flagellum at each pole

c) Lophotrichate: tuff flagella at one or both poles.

d) Peritrichate: flagella completely surrounding the

bacterial cell.

The flagella are too thin to be seen in ordinary

stained preparations, but become visible only in silver

impregnated preparation or in electron microscopy.

The functions of flagella are:

1. motility (mainly)

2. attachment to site of infection e.g. stomach.

3. Diversity of antigens, this is used for diagnosis and

this is mainly in Salmonella typhi (the causative agent

of typhoid fever).

4. Invasion.

5. Colonization (form colony).

3- Pili:Extended from cytoplasmic membrane, they

are shorter and finer filaments than the flagella. These are the pili. They are made of protein

(pilin).Pili are divided into two groups:

a. Sex pili, referred to as sex pili because of their role during conjugation when genes are

transferred from one cell(donor) to another cell (recipient).

b. Common pili, or Fimbriae: these are thought to be the organs of adhesion that help bacteria to

attach to the host cells (Virulent Factor).

4- Spores:-

These are dense structure produced by some

bacteria, e.g. the Bacilli and Clostridia groups, that enable

them to survive adverse environmental conditions. They

develop within and at the expense of the

vegetative cell. The spore comprises the

chromosomal material surrounded by several

walls layers.

Chemically, endospore has a large amount of Ca++ and less

number of enzymes. Dpiclonic acid is also presents in spore.

Spores are resistant to heat, stains, desiccation,

chemicals and disinfectants.

Each spore germinates to produce a vegetative cell during

favorable conditions.

The location and shape of the spore in the cell may be of

diagnostic assistance, e.g. the spores of Clostridium

tetani are terminal, and the diameter is greater than that of

the parent cell, so that they are characteristically of drum

stick appearance.The positions of the spores are described as:Terminal , Sub terminal , or Central

The shapes of spores are described as:Ovoid (Oval) , Spherical , Cylindrical .

Classification of bacteria:1- phenotypic classification: The microscopic and macroscopic morphologies of bacteria were the first

characteristic used to identify bacteria. The phenotypic classification include:

* Microscopic morphology (cocci, bacilli, curved, spiral, spirochetes and Vibrio), also the ability to retain the gram stain

into gram positive and gram negative. * Macroscopic morphology (appearance of colonies of

bacteria on culture media (colonies size, shape, pigment production and reaction on media.

* Biotyping (using biochemical test or reaction) *Serotyping (by detection bacterial antigens using

serological test. * phage typing (susceptibility to viruses that infect bacteria)

2- Analytic classification: using chromatographic pattern include:

* Cell wall fatty acid analysis * Whole cell lipid analysis

* Whole cell protein analysis * Multilocus enzyme electrophoresis

3- Genotypic classification: the most precise method for classifying bacteria is by analysis by their genetic material that

include: * Guanine plus cytosine ratio

* DNA hybridization * Nucleic acid sequence analysis

* plasmid analysis * chromosomal DNA fragments

* Ribotyping

•Others useful classification:* According to their oxygen requirement into:

1- Aerobic 2- Anaerobic 3- Facultative anaerobic

4- Microaerophilic.* According to temperature into:

1- Thermophilic 2- Mesophilic 3- Psychrophilic* According to their growth factors into:

1- Fastidious organism (required special growth factor)

2- non fastidious (simple)

Cell wall of gram positive bacteriaCell wall of gram positive bacteria

Bacterial terminal spores (Bacterial terminal spores (Clostridium tetaniClostridium tetani))

Bacterial flagellumBacterial flagellum