MICROBIOLOGY

Microbiology, study of microorganisms, or microbes, a diverse group of generally minute, simple

life-forms that include bacteria, archaea, algae, fungi, protozoa, and viruses. The field is concerned
with the structure, function, and classification of such organisms and with ways of both exploiting
and controlling their activities.

CLINICAL MICROBILOGY

Clinical Microbiology is a branch of medical science concerned with the prevention, diagnosis and
treatment of infectious diseases. Moreover, this field of science is concerned about various clinical
applications of microbes for the improvement of health. There are four kinds of microorganisms that
cause infectious disease: bacteria, fungi, parasites and viruses.

Clinical microbiology is a discipline that encompasses a broad range of testing methodologies, and it
is complex in terms of organisms and methods used to isolate and identify them. Although significant
improvements in testing methodologies have been made, clinical microbiology remains heavily
reliant on culture-based methods and phenotypic methods for identification of culture organisms.

The wide variety of pathogens and testing methods that are available makes microbiological testing
challenging, and thus error detection and correction are important components of quality
microbiology laboratory testing. Errors may occur at all stages of testing (pre-analytical, analytical,
and post-analytical), and an error in one stage of testing is likely to overlap with or lead to errors in
other stages (e.g., incorrect specimen collection can lead to culture, identification, and reporting of
organisms that are not involved in the disease process and to incorrect or
unnecessary antimicrobial therapy as a result).

PREPARATION OF SAMPLES FOR ELECTRON MICROSCOPE

Electron microscopes are very powerful tools for visualising biological samples. They enable
scientists to view cells, tissues and small organisms in very great detail. However, these biological
samples can’t be viewed on electron microscopes whilst alive. Instead, the samples must undergo
complex preparation steps to help them withstand the environment inside the microscope.

The preparation process kills the tissue and can also cause changes in the sample’s appearance.For
scientists who wish to view biological samples, this poses a challenge – how can the sample be
preserved so that it looks as much as possible like it would in the living organism, while still being
able to withstand being visualised in the electron microscope.

Surviving hostile environment

There are two reasons why living things can’t survive in an electron microscope:

• The power of the electron beam that’s directed at the sample.

• The vacuum inside the microscope.

The electron beam inside a transmission electron microscope (TEM) causes problems for biological
samples because of its high energy. It needs to have enough energy to pass right through the sample and
out the other side. The temperature can get up to 150°C where the beam hits the sample. This temperature
is far too high for living cells to survive. Scanning electron microscopes (SEMs) use a lower-energy
electron beam, but it can still be damaging to the sample.

The vacuum inside an electron microscope is important for its function. Without a vacuum, electrons
being aimed at the sample would be deflected (knocked off course) when they hit  air particles. But liquid
water, which is abundant in biological samples, evaporates immediately in a vacuum. If this happened, a
biological sample would vaporise in front of your eyes!

To be visualised by an electron microscope, biological samples need to be:

• fixed (stabilised) so the electron beam doesn’t destroy them

• dried thoroughly so the vacuum doesn’t affect them.

Fixation: a snapshot of the living sample

The first – and perhaps most important – step in the preparation process is fixation. In this step, living
tissue is chemically treated to stabilise it. This kills the tissue sample at the same time. It’s important
to fix a sample as quickly as possible because, as soon as tissue is removed from its natural
environment, it starts to change. For instance, oxygen levels start to drop as soon as tissue is removed
from an organism. This causes mitochondria to start to change their appearance. Another common
change in the fixation process is that lipids tend to form micelles.

Looking out for artefacts of fixation

Micelles and strange-shaped mitochondria are examples of artefacts – structures that are seen under
the microscope but aren’t found in living cells. It’s very important to be aware that artefacts can be
introduced during fixation so that you don’t mistake them for real parts of your sample. Telling the
difference between an artefact and a ‘real’ structure can be difficult. To minimise the introduction of
artefacts, scientists are continually experimenting with new ways to prepare samples. One approach is
to freeze the sample very quickly instead of fixing it. Providing the sample stays cold enough, this
‘locks up’ the water and prevents it from evaporating inside the microscope.

Freezing samples is common in SEM (and is known as cryoSEM). It is still in the early stages of
development for TEM .Electron microscopy is a valuable tool used to obtain high-resolution images
in a variety of applications, including biomedical research, forensics, and technology. Electron
microscopes can capture much higher resolution images than light microscopes, contributing
information that is otherwise unattainable.

Every electron microscope works by accelerating a focused stream of electrons in a vacuum towards a
sample. Interactions between the electron beam and the sample create an image, similar to how optical
microscopes use light to capture images. The image created reveals details of a sample’s surface or
internal composition, depending on the type of electron microscope that is used.

Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM) are the two most
common types of electron microscopy. TEM and SEM differ in how they work and what types of images
they are able to capture. This article will overview SEM and TEM, including what they are, how they
work, and how they compare to one another.

What is SEM? And How does SEM work?

SEM can stand for either Scanning Electron Microscopy or Scanning Electron Microscope. An SEM
is a kind of electron microscope that uses a fine beam of focused electrons to scan a sample’s surface.
The microscope records information about the interaction between the electrons and the sample,
creating a magnified image. SEM has the potential to magnify an image up to 2 million times.

SEM images give insight into a sample’s topography and elemental composition. SEM is able to
capture 3-D black-and-white images of thin or thick samples. The sample’s size is limited only by the
size of the electron microscope chamber.

Working

To obtain a high-resolution image, an electron source (also known as an electron gun) emits a stream
of high-energy electrons towards a sample. The electron beam is focused using electromagnetic
lenses. Once the focused stream reaches the sample, it scans its surface in a rectangular raster.

The interaction between the electron beam and the sample creates secondary electrons, backscattered
electrons, and X-rays. These interactions are captured to create a magnified image.
TEM?

TEM can stand for Transmission Electron Microscopy or Transmission Electron Microscope (TEM).
A TEM is a type of electron microscope that uses a broad beam of electrons to create an image of a
sample’s internal structure. A beam of electrons is transmitted through a sample, creating an image
that details a sample’s morphology, composition, and crystal structure.

Samples must be incredibly thin, often less than 150 nm thick, to allow electrons to pass through
them. After the transmission of the electrons through the sample, they arrive at a detector below and a
2-D image is created.
 TEMs have an incredible magnification potential of 10-50 million times. The details provided are at
the atomic level, the highest resolution of any electron microscope. TEMs are often used to examine
molecular and cellular structures.

How does a TEM work?

An electron source sends a beam of electrons through an ultrathin sample. When the electrons
penetrate the sample, they pass through lenses below. This data is used to create images directly on a
fluorescent screen or onto a computer screen using a charge-coupled device (CCD) camera.

SEM vs TEM advantages

Scanning Electron Microscopes and Transmission Electron Microscopes each contain unique advantages
when compared to the other.

In comparison to TEMs, SEMs:

 • Cost less

• Take less time to create an image

• Require less sample preparation

• Accept thicker samples

• Can examine larger samples

In comparison to SEMs, TEMs:

• Create higher resolution images

• Provide crystallographic and atomic data

• Create 2-D images that are often easier to interpret than SEM 3-D images

• Allow users to examine more characteristics of a sample

SEM vs TEM similarities and differences

Scanning Electron Microscopes Transmission Electron Microscopes (TEM)

(SEM)

Electron stream Fine, focused beam Broad beam

Image taken Topographical/surface Internal structure

Resolution Lower resolution Higher resolution

Magnification Up to 2,000,000 times  Up to 50,000,000 times

Image dimension 3-D 2-D

Sample thickness Thin and thick samples okay Ultrathin samples only

Penetrates sample No Yes

Scanning Electron Microscopes Transmission Electron Microscopes (TEM)

(SEM)
Sample restriction Less restrictive More restrictive

Sample preparation Less preparation required More preparation required

Cost Less expensive More expensive

Speed Faster Slower

Operation Easy to use More complicated; requires training

STAINING METHODS

Staining
Staining is technique used in microscopy to enhance contrast in the microscopic
image. Stains and dyes are frequently used in biological tissues for viewing,
often with the aid of different microscopes. Stains may be used to define and
examine bulk tissues (highlighting, for example, muscle fibers or connective
tissue), cell populations (classifying different blood cells, for instance), or
organelles within individual cells.

Bacteria have nearly the same refractive index as water, therefore, when they
are observed under a microscope they are opaque or nearly invisible to the
naked eye.

Different types of staining methods are used to make the cells and their internal
structures more visible under the light microscope. Microscopes are of little use
unless the specimens for viewing are prepared properly. Microorganisms must
be fixed & stained to increase visibility, accentuate specific morphological
features, and preserve them for future use.

Stain

A stain is a substance that adheres to a cell, giving the cell color. The presence
of color gives the cells significant contrast so they are much more visible.
Different stains have different affinities for different organisms, or different
parts of organisms. They are used to differentiate different types of organisms
or to view specific parts of organisms.

Staining techniques

• Direct staining - The organism is stained and background is left unstained

• Negative staining - The background is stained and the organism is left

unaltered

Stains are classified as

• Simple stain

• Differential stain
 • Structural or special stains

Fixing Before staining it is essential to fix the bacterial sample on to the slide.
Smear is prepared in the following way:

• With a wire loop place a small drop of the broth culture or a loop full of
bacteria on a clean slide.
• Place a drop of water over it.
• Spread the culture so as to form a thin film.
• Allow slide to dry in the air or by holding it above a bunsen flame.
• Avoid excess heating.

The purpose of fixation is to kill the microorganisms, coagulate the protoplasm

of the cell and cause it to adhere to the slide Simple Staining. The staining
process involves immersing the sample (before or after fixation and mounting)
in dye solution, followed by rinsing and observation.

Many dyes, however, require the use of a mordant, a chemical compound that
reacts with the stain to form an insoluble, coloured precipitate. When excess
dye solution is washed away, the mordant stain remains. Simple staining is one
step method using only one dye.

Basic dyes are used in direct stain and acidic dye is used in negative stain.
Simple staining techniques is used to study the morphology better, to show the
nature of the cellular contents of the exudates and also to study the intracellular
location of the bacteria.

Commonly used simple stains are

– Methylene blue

– Dilute carbol fuchsin

Polychrome methylene blue

Simple Staining Procedure:

 When a single staining-reagent is used and all cells and their structures stain in
the same manner, the procedure is called simple staining procedure. This
procedure is of two types – positive and negative. In positive staining, the stain
(e.g., methylene blue) is basic (cationic) having positive charge and attaches to
the surface of object that is negatively charged.

In negative staining, the stain (e.g., India ink, nigrosin) is acidic (anionic)
having negative charge and is repelled by the object that is negatively charged,
and thus fills the spaces between the objects resulting in indirect staining of the
object.

GRAM STAINING
This differential staining procedure separates most bacteria into two groups on the basis of cell
wall composition:

• Gram-positive bacteria (thick layer of peptidoglycan-90% of cell wall)- stains purple

• Gram-negative bacteria (thin layer of peptidoglycan-10% of cell wall and high lipid

content) –stains red/pink

Steps of Gram Staining

• Fixation of clinical materials to the surface of the microscope slide either by heating or
by using methanol. (# Methanol fixation preserves the morphology of host cells, as well
as bacteria, and is especially useful for examining bloody specimen material).

• Application of the primary stain (crystal violet). Crystal violet stains all cells
blue/purple

• Application of mordant: The iodine solution (mordant) is added to form a crystal violet-

iodine (CV-I) complex; all cells continue to appear blue.

• Decolorization step: The decolorization step distinguishes gram-positive from gram-

negative cells.

• The organic solvent such as acetone or ethanol extracts the blue dye complex from the
lipid-rich, thin-walled gram-negative bacteria to a greater degree than from the lipid-
poor, thick-walled, gram-positive bacteria.  The gram-negative bacteria appear colorless
and gram-positive bacteria remain blue.

• Application of counterstain (safranin): The red dye safranin stains the decolorized

gram-negative cells red/pink; the gram-positive bacteria remain blue.
Gram Staining Procedure

• Flood air-dried, heat-fixed smear of cells for 1 minute with crystal violet staining reagent.
Please note that the quality of the smear (too heavy or too light cell concentration) will
affect the Gram Stain results.

• Wash slide in a gentle and indirect stream of tap water for 2 seconds.

• Flood slide with the mordant: Gram’s iodine. Wait 1 minute.

 • Wash slide in a gentle and indirect stream of tap water for 2 seconds.

• Flood slide with decolorizing agent (Acetone-alcohol decolorizer). Wait 10-15 seconds or

add drop by drop to slide until decolorizing agent running from the slide runs clear.

• Flood slide with a counterstain, safranin. Wait 30 seconds to 1 minute.

• Wash slide in a gentile and indirect stream of tap water until no color appears in the
effluent and then blot dry with absorbent paper.

• Observe the results of the staining procedure under oil immersion (100x) using a Bright
field microscope.

Ziehl-Neelsen technique (AFB Staining)

Ziehl-Neelsen (ZN) method of Acid Fast staining technique is used to

stain Mycobacterium species including M. tuberculosis, M. ulcerans, and M.
leprae and nontuberculous mycobacteria (NTM).  Detection of acid-fast bacilli (AFB) in
stained and acid-washed smears examined microscopically may provide the initial
bacteriologic evidence of the presence of mycobacteria in a clinical specimen. Smear
microscopy is the quickest and easiest procedure that can be performed.

Principle of Ziehl-Neelsen method of acid-fast staining

Mycobacteria, which do not stain well by Gram stain, are stained with carbol fuchsin combined
with phenol.\

• In the ‘hot’ ZN technique, the phenol-carbol fuchsin stain is heated to enable the dye to
penetrate the waxy mycobacterial cell wall.

• In the ‘cold’ technique known as Kinyoun Method, stain are not heated but the
penetration is achieved by increasing concentration of basic fuchsin and phenol and
incorporating a ‘wetting agent’ chemical.

The stain binds to the mycolic acid in the mycobacterial cell wall. After staining, an acid
decolorizing solution is applied. This removes the red dye from the background cells, tissue
fibres, and any organisms in the smear except mycobacteria which retain (hold fast to) the dye
and are therefore referred to as acid fast bacilli (AFB).

• Following decolorization, sputum smear is counterstained with malachite green, or

methylene blue which stains the background material, providing a contrast colour against
which the red AFB can be seen.

• Among the Mycobacterium species, M. tuberculosis and M. ulcerans are strongly acid

fast. When staining specimens for these species, a 3% v/v acid alcohol is used to
 decolorize the smear, where as M. leprae is only weakly acid fast. 0.5-1% v/v
decolorizing solution is therefore used for M. leprae smears and also different staining
and decolorizing time.

Sample Collection & Preparation : Due to overnight accumulation of secretions, first morning
specimens are more likely to yield better recovery of AFB.

• Direct Smear: Smear prepared directly from a patient specimen prior to processing.

• Indirect Smear:  Smear prepared from a processed specimen after centrifugation (is used

to concentrate the material)

Reagents required:

• Carbol fuchsin stain (filtered)

• Acid alcohol 3% v/v (or 20% sulfuric acid)

• Malachite green 5 g/l (0.5% w/v) or Methylene blue, 5g/l

STRUCTURE OF BACTERIA AND VIRUS

Structure of Bacteria

Bacteria are unicellular organisms belonging to the prokaryotic group where the organisms
lack a few organelles and a true nucleus”.The bacteria diagram given below represents the
structure of bacteria with its different parts.  The cell wall, plasmid, cytoplasm and flagella
are clearly marked in the diagram.

The structure of bacteria is known for its simple body design. Bacteria are single-celled
microorganisms with the absence of the nucleus and other cell organelles; hence, they are
classified as prokaryotic organisms.They are also very versatile organisms, surviving in
extremely inhospitable conditions. Such organisms are called extremophiles

Extremophiles are further categorized into various types based on the types of environments
they inhabit:

• Thermophiles

• Acidophiles

• Alkaliphiles

• Osmophiles

• Barophiles

Cryophiles
Another fascinating feature of bacteria is their protective cell wall, which is made up of a
special protein called peptidoglycan. This particular protein isn’t found anywhere else in
nature except in the cell walls of bacteria.But few of them are devoid of this cell wall, and
others have a third protection layer called capsule. On the outer layer, one or more flagella or
pili is attached, and it functions as a locomotory organ.

Pili can also help certain bacteria to attach themselves to the host’s cells. They do not contain
any cell organelle as in animal or plant cell except for ribosomes.Ribosomes are the sites of
protein synthesis. In addition to this DNA, they have an extra circular DNA called plasmid.
These plasmids make some strains of bacteria resistant to antibiotics.

Structure of Virus

Viruses are infectious agents that replicate inside the body of a host.Viruses are non-cellular,
microscopic infectious agents that can only replicate inside a host cell. From a biological
perspective, viruses cannot be classified either a living organism or non-living. This is due to
the fact that they possess certain defining characteristic features of living organisms and non-
living entities.In a nutshell, a virus is a non-cellular, infectious entity made up of genetic
material and protein that can invade and reproduce only within the living cells of bacteria,
plants and animals.

For instance, a virus cannot replicate itself outside the host cell. This is because viruses
lack the required cellular machinery. Therefore, it enters and attaches itself to a specific host
cell, injects its genetic material, reproduces by using the host genetic material and finally the
host cell splits open, releasing the new viruses.Viruses can also be crystallized, which no
other living organisms can do. It is these factors that lead to viruses being classified in the
grey area – between the living and non-living.
Bacteriophage and HIV

These microbes belong to the family viridae and genus virus.  Viruses could not be placed in
any of the kingdoms because they are practically neither living nor dead. Once a susceptible
cell is infected, a virus can start the cell machinery to generate more virus. Viruses are
composed of a core of DNA or RNA surrounded by a protein coat. They are very small and
their size ranges from 20 nanometers to 250 nanometers. Therefore, they can only be seen
with an electron microscope

Many viruses have either DNA or RNA as the genetic element and the nucleic acid with
single or double strands. The whole infectious virus, called as virion has nucleic acid and an
outer shell of proteins. The simplest virus includes DNA or RNA for encoding four proteins
and the most complex encodes 100-200 proteins.The study of viruses is called as virology.
ROUTES OF INFECTION AND INFECTION SPREAD

The Main Routes Infection can Enter the Body are:

• Body fluids – A body fluid e.g. blood, urine, pus, saliva from one person enters the body
of another e.g. through cuts. Through saliva (e.g. glandular fever). Through contaminated
substances entering the bloodstream e.g. tetanus can be contracted through wounds when
handling contaminated soil.

• Through the air – e.g. tuberculosis. Droplet infections are carried through the air
through coughs and sneezes (e.g. common cold, respiratory viruses).

• Through touch – Those caught by directly touching the skin/skin to skin contact (these
are often referred to as contagious) of someone who has a particular kind of infection
such as chickenpox.

• Touching an infected object which an infected person has touched e.g. door handles,
paper and cooking implements – e.g. Norovirus (winter vomiting infection) can be caught
this way.

• Through ingestion – contaminated food causes food poisoning (e.g. salmonella and
campylobacter) or water (e.g. cholera)

• Through bites from other creatures – e.g. infections from dog and cat bites, (e.g. blood
poisoning/sepsis) or through parasites e.g. insect bites/stings (e.g. malaria).

• Insect bites and stings can also cause allergic reactions which are not due to infections
but due to toxins which exist in the venom which has entered the body.

Routes of Infection spread

• Infection control and prevention depends on disrupting the transmission of

pathogens from their source (the infected animal or human) to new hosts (animal or
human) or locations.

• Understanding routes of disease transmission and how it contributes to the spread

of organisms allows for the identification of effective prevention and control
measures not only for specific diseases, but also other pathogens transmitted by a
similar route, including unanticipated infectious diseases.

• The transmission of microorganisms can be divided into the following five main
routes: direct contact, fomites, aerosol (airborne), oral (ingestion), and vectorborne. 

• Some microorganisms can be transmitted by more than one route.

Direct Contact Transmission

Direct contact transmission occurs through direct body contact with the tissues or fluids of an
infected individual. Physical transfer and entry of microorganisms occurs through mucous
membranes (e.g., eyes, mouth), open wounds, or abraded skin.

Direct inoculation can occur from bites or scratches. Examples include organisms such as
rabies, Microsporum, . This is probably the most common and highest-risk route of pathogen
transmission to patients and personnel.

•  

Fomite Transmission

Fomite transmission involves inanimate objects contaminated by an infected individual that

then come in contact with a susceptible animal or human. Fomites can include a wide variety
of objects such as exam tables, cages, kennels, medical equipment, environmental surfaces,
and clothing. Disease examples include canine parvovirus and feline calicivirus infections.
Aerosol (Airborne) Transmission

Aerosol transmission encompasses the transfer of pathogens via very small particles or
droplet nuclei. Aerosol particles may be inhaled by a susceptible host or deposited onto
mucous membranes or environmental surfaces. This can occur from breathing, coughing,
sneezing, or vocalization of an infected individual, but also during certain medical
procedures).

Very small particles may remain suspended in the air for extended periods and be
disseminated by air currents in a room or through a facility.However, most pathogens
pertinent to companion animal veterinary medicine do not survive in the environment for
extended periods or do not travel great distances due to size and as a result require close
proximity or contact for disease transmission.

Oral (Ingestion) Transmission

The ingestion of pathogenic organisms can occur from contaminated food or water as well as
by licking or chewing on contaminated objects or surfaces. Environmental contamination is
most commonly due to exudates, feces, urine, or saliva. Examples of diseases acquired via
oral transmission include feline panleukopenia and infections caused
by Campylobacter, Salmonella, Escherichia coli, and Leptospira.

Vector-Borne Transmission

Vectors are living organisms that can transfer pathogenic microorganisms to other animals or
locations and include arthropod vectors (e.g., mosquitoes, fleas, ticks) and rodents or other
vermin. Vector-borne transmission can be an important route of transmission in climates
where these pests exist year-round and may be brought into the practice by an infested
patient. Examples of vector-borne diseases include heartworm disease, Bartonella infection,
Lyme disease (borreliosis), and plague.
ENDOGENOUS INFECTIONS AND EXOGENOUS INFECTIONS

Endogenous Infections

In the case of endogenous infections, we become infected with our own bacteria, our own
microflora.This might happen if a barrier between sterile and non-sterile tissues is broken,
such as with a bowel perforation.A patient with a compromised immune system, such as after
chemotherapy, may become sick from a bacteria already present in his/her body that grows
unchecked.A dormant pathogen might also become reactivated and infect the host, such as in
the case of tuberculosis.

From an infection prevention standpoint, avoiding endogenous infections focuses on careful

protocols during invasive procedures, antibiotic stewardship, and working to keep the
patient's immune system working as well as possible.

Exogenous Infections

Exogenous infections, in contrast, involve a pathogen entering a patient's body from

environment.These pathogens can be introduced through a contaminated device, healthcare
worker, surface, or other vector.Patients with open incisions, indewelling devices, and
compromised immune systems are especially at risk for exogenous infections.

Eliminating exogenous infections focuses almost entirely on reducing the bioburden in the
patient's environment. If the reservoirs for pathogens can be reduced, the opportunities for
cross-contamination diminish.Everyone agrees that hand hygiene is the single most important
way to reduce transmission of pathogens to vulnerable patients.Other interventions focus on
environmental decontamination of surfaces and other fomites, including daily cleaning, UV
light, and self-sanitizing surfaces.

Whether exogenous or endogenous, bacteria needs to stay where it's supposed to be. Skin
flora needs to stay on the skin, not get into the respiratory tract. Bacteria that help us digest
food need to stay in the gut, not get into our food. Equally important, bacteria in our
environment needs to stay off our hands, our devices, and our patients. It falls to all of us to
play traffic controller in this microbiologic universe to give ourselves and others the best
chance to stay healthy.
CULTURE MEDIA AND ITS TYPES

Culture media contains nutrients and physical growth parameters necessary for microbial
growth. All microorganisms cannot grow in a single culture medium and in fact, many can’t
grow in any known culture medium. Culture medium or growth medium is a liquid or gel
designed to support the growth of microorganisms. There are different types of media
suitable for growing different types of cells.

Microbiological cultures used for growing microbes

Nutrient broths and agar plates

These are the most common growth media, although specialized media are sometimes
required for microorganism and cell culture growth. Some organisms, termed fastidious
organisms, need specialized environments due to complex nutritional requirements. Viruses,
for example, are obligate intracellular parasites and require a growth medium containing
living cells. Many human microbial pathogens also require the use of human cells or cell
lysates to grow on a media.

The most common growth media nutrient broths (liquid nutrient medium) or LB medium
(Lysogeny Broth) are liquid. These are often mixed with agar and poured into Petri dishes to
solidify. These agar plates provide a solid medium on which microbes may be cultured. They
remain solid, as very few bacteria are able to decompose agar. Many microbes can also be
grown in liquid cultures comprised of liquid nutrient media without agar.

Defined vs Undefined media

• A defined medium will have known quantities of all ingredients. For microorganisms, it

provides trace elements and vitamins required by the microbe and especially a defined
carbon and nitrogen source. Glucose or glycerol are often used as carbon sources, and
ammonium salts or nitrates as inorganic nitrogen sources.

• An undefined medium has some complex ingredients, such as yeast extract, which

consists of a mixture of many, many chemical species in unknown proportions.
Undefined media are sometimes chosen based on price and sometimes by necessity –
some microorganisms have never been cultured on defined media.
Common broadly-defined culture media

Nutrient media – A source of amino acids and nitrogen (e.g., beef, yeast extract).

This is an undefined medium because the amino acid source contains a variety of compounds
with the exact composition being unknown. These media contain all the elements that most
bacteria need for growth and are non-selective, so they are used for the general cultivation
and maintenance of bacteria kept in laboratory-culture collections.

Minimal media

Media that contains the minimum nutrients possible for colony growth, generally without the
presence of amino acids, and are often used by microbiologists and geneticists to grow “wild
type” microorganisms.These media can also be used to select for or against the growth of
specific microbes. Usually a fair amount of information must be known about the microbe to
determine its minimal media requirements.

Selective media 

Used for the growth of only selected microorganisms. For example, if a microorganism is
resistant to a certain antibiotic, such as ampicillin or tetracycline, then that antibiotic can be
added to the medium in order to prevent other cells, which do not possess the resistance,
from growing.
Differential media 

Also known as indicator media, are used to distinguish one microorganism type from
another growing on the same media. This type of media uses the biochemical characteristics
of a microorganism growing in the presence of specific nutrients or indicators added to the
medium to visibly indicate the defining characteristics of a microorganism. This type of
media is used for the detection and identification of microorganisms.

Growth curve

Growth curve, in biology, a curve in graph form that shows the change in the number of
cells (or single-celled organisms) in an experimental culture at different times. Growth
curves are also common tools in ecological studies; they are used to track the rise and fall of
populations of plants, animals, and other multicellular organisms over time.
 DISEASES CAUSED BY BACTERIA,FUNGI,PROTOZOAL,VIRUS AND
HELMINTHES

Bacterial Diseases

Bacterial diseases include any type of illness caused by bacteria. Bacteria are a type of
microorganism, which are tiny forms of life that can only be seen with a microscope. Other
types of microorganisms include viruses, some fungi, and some parasites.Millions of bacteria
normally live on the skin, in the intestines, and on the genitalia. The vast majority of bacteria
do not cause disease, and many bacteria are actually helpful and even necessary for good
health. These bacteria are sometimes referred to as “good bacteria” or “healthy bacteria.”

Harmful bacteria that cause bacterial infections and disease are called pathogenic bacteria.
Bacterial diseases occur when pathogenic bacteria get into the body and begin to reproduce
and crowd out healthy bacteria, or to grow in tissues that are normally sterile. Harmful
bacteria may also emit toxins that damage the body.

Common pathogenic bacteria and the types of bacterial diseases they cause include:

• Escherichia coli and Salmonella cause food poisoning.

• Helicobacter pylori cause gastritis and ulcers.

• Neisseria gonorrhoeae causes the sexually transmitted disease gonorrhea.  

• Neisseria meningitidis causes meningitis.

• Staphylococcus aureus causes a variety of infections in the body, including

boils, cellulitis, abscesses, wound infections, toxic shock syndrome, pneumonia, and food
poisoning.

• Streptococcal bacteria cause a variety of infections in the body, including pneumonia,

meningitis, ear infections, and strep throat.

Fungal infections

Fungal infections are common throughout much of the natural world. In humans, fungal
infections occur when an invading fungus takes over an area of the body and is too much for
the immune system to handle.Fungi can live in the air, soil, water, and plants. There are also
some fungi that live naturally in the human body.

Like many microbes, there are helpful fungi and harmful fungi. When harmful fungi invade the
body, they can be difficult to kill, as they can survive in the environment and re-infect the person
trying to get better.
The symptoms of a fungal infection will depend on the type, but common symptoms include the
following:

• skin changes, including red and possibly cracking or peeling skin

itching

Athlete’s foot

Tinea pedis or athlete’s foot is a common fungal infection that affects the foot.The fungus
grows perfectly in warm, moist environments, such as socks and shoes, sports equipment,
and locker rooms.

Yeast infection

Vaginal yeast infections are a common form of Candida overgrowth in women, usually

caused by Candida albicans.An overgrowth of Candida disrupts the normal balance of the
bacteria and yeast in the vagina. This imbalance of bacteria may be due to antibiotics, stress,
and hormone imbalances, or poor eating habits, among other things.Candida infections can
also commonly cause fungal toenail infections and diaper rash.

Jock itch

Tinea cruris, commonly known as jock itch, is another common fungal skin infection.These
fungi love warm and damp environments and thrive in moist areas of the body, such as the
groin, buttocks, and inner thighs. Jock itch may be more common in summer or in warm,
humid areas of the world.Jock itch is mildly contagious and is often spread through direct
contact with an infected person or an object that is carrying the fungus.

Ringworm

Tinea corporis or ringworm is a skin infection caused by a fungus that lives on dead tissues,
such as the skin, hair, and nails. Ringworm is the fungus that causes both jock itch and
athlete’s foot. When it appears anywhere else on the body, the infection is just called
ringworm.

Dieases by Protozoal
Trypanosoma Protozoa

Members of the genus Trypanosoma are flagellate protozoa that cause sleeping sickness.

The parasites are spread by insect vectors.Trypanosoma parasites enter a person’s blood
when the vector bites. Then they spread to other tissues and organs.The diseases may be fatal
without medical treatment.

Giardia Protozoa

Giardia are flagellate protozoa that cause giardiasis. The parasites enter the body through
food or water that has been contaminated by feces of infected people or animals. The
protozoa attach to the lining of the host’s small intestine, where they prevent the host from
fully absorbing nutrients. They may also cause diarrhea, abdominal pain, and fever. A picture
of a Giardia protozoan opens this concept.

Plasmodium Protozoa

Plasmodium protozoa cause malaria.The parasites are spread by a mosquito vector. Parasites

enter a host’s blood through the bite of an infected mosquito. The parasites infect the host’s
red blood cells, causing symptoms such as fever, joint pain, anemia, and fatigue.

Dieases by Helminthiasis

Helminthiasis, also known as worm infection, is any macroparasitic disease of humans and

other animals in which a part of the body is infected with parasitic worms, known
as helminths. There are numerous species of these parasites, which are broadly classified
into tapeworms, flukes, and roundworms.They often live in the gastrointestinal tract of
their hosts, but they may also burrow into other organs, where they induce physiological
damage.