Microbiology and Techniques | Study

Running head: BIOLOGY
MICROBIOLOGY AND TECHNIQUES
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Running head: BIOLOGY1
Table of Contents
First section................................................................................................................................2
Preparation of growth media:.................................................................................................2
Inoculation of growth media:.................................................................................................3
Second Section...........................................................................................................................4
Factors that influence microbial growth:...............................................................................4
Effect of temperature on microbial growth (Fig 2):...............................................................5
Measuring microbial growth (Haemocytometer and Colorimeter):.......................................6
Nutritional requirements of bacteria......................................................................................7
Third Section..............................................................................................................................8
Effects of antibiotics and antiseptics:.....................................................................................8
Use of biocontainment in microbiological laboratories:........................................................9
References................................................................................................................................11

Running head: BIOLOGY2
First section
Preparation of growth media:
Microbiology is a vast field of study which involves the characterization of various
organisms that are invisible to the naked eye. These microorganisms live freely in the
environment, however, most of them require a substratum to grow and survive. The
substratum should contain the nutrient materials required for the growth of that organism. In
vitro growth requires the use of specialized media known as the growth media. Most of these
media are obtained in the form of powders. Therefore they are mixed with a semi-solid
substance (agar) to provide a nutritious substratum for the growth of microorganisms. The
most significant growth media for all the microorganisms are liquid nutrient media (nutrient
broth) also known as LB. Growth media can be of three types liquid, solid and semisolid
which supports the microorganism growth in vitro. Liquid media is prepared by mixing the
nutrient powdered with molten agar followed by a volume makeup with distilled water. These
media are stored either in the conical flask or test tubes and sterilized in an autoclave (1). The
main components of growth media are various growth factors and cell building biomolecules
such as protein, carbohydrates, and lipids in specified amounts. Culture media belongs to the
first category of growth media with specified amounts of carbon sources, various salts,
nitrogen, and amino acid sources and water. This is the compositional preparation of an
undefined media. A defined medium includes known chemicals and also lacks the presence
of yeast, plant or animal tissues inside the media. Examples of nutrient media include nutrient
agar and trypticase soy agar. Minimal media is a type of defined media in which the various
types of microorganisms are grown and selected on the media (2). This media has also been
used for recombinant selection study. Selective media is the third category of growth media
which is used in microbiology. Eosin methylene blue media includes dyes, which help in the

Running head: BIOLOGY3
selection of coliforms and other gram-negative bacteria since it is toxic to the gram-positive
bacteria. Eosin methylene blue (EMB) media consists of eosin and methylene blue in a 6:1
ratio. For example, E.coli growth on EMB media produces a distinctive metallic green sheen
which helps in the direct identification of the organism. Differential media and enriched
media are the two contrasting types of media. Enriched media is prepared by almost every
kind of nutrients required to grow various kinds of microorganisms. Chocolate agar and
blood agar are the most significant sources of growth media widely used in microbiology (3).
Inoculation of growth media:
Inoculation is the process of the addition of bacterial culture on the growth media with
the help of an inoculation loop. The inoculation loop is inserted into the mother culture and
one loop full of culture is transferred to either a test tube broth media or agar media in a Petri
plate. There are three types of inoculation mechanisms for plate media named as streaking,
spreading and pour plate method (4). From all these techniques streak plate procedure is used
in most of the cases for single colony isolation. This technique is time-consuming and allows
proper colony formation. There are two types of streak plate method, one is continuous and
another is discontinuous. Both of these types involve streaking the bacterial culture on a Petri
plate by using an inoculation needle or loop. The streaking process can be used for slant
cultures also since the agar media is slanted inside a culture tube and then solidified. Slant
preparation is required for the storage of a particular culture of bacteria for further use. When
the bacteria are required to be used in the future for further testing, a single colony is isolated
from the slant cultures and then transferred to a Petri plate. Lawn preparation is done by
spread plate technique or pour plate technique (5). This process requires the pouring of a
liquid culture of bacterial evenly on a plate with nutrient agar media. After the incubation is
done at the specified temperature, the colonies become randomly scattered around the plate
forming a lawn of single microbial colonies. Spread plate or pour plate (lawn) methods are