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Gebze Technical
University
Microbiology Laboratory – Report 5
Turkey
Ebru AKHARMAN
20.04.2018
The Culturing Microorganisms
AIM
In this experiment, 4 types of media are learned and inoculations are performed on these medyums.
Numerous special-purpose media are available for functions including the following:
1. Isolation of bacterial types from a mixed population of organisms.
2. Differentiation among closely related groups of bacteria on the basis of macroscopic appearance of the
colonies and biochemical reactions with in the medium.
3. Enumeration of bacteria in sanitary microbiology, such as in water and sewage, and also in food and dairy
products.
4. Assay of naturally occurring substances such as antibiotics, vitamins, and products of industrial
fermentation.
5. Characterization and identification of bacteria by their abilities to produce chemical changes in different
media.
In addition to nutrients necessary for the growth of all bacteria, special-purpose media contain both nutrients
and chemical compounds important for specific metabolic pathways in different types of bacteria.
1. Solid medium
solid medium contains agar at a concentration of 1.5-2.0% or some other, mostly inert solidifying agent.
Solid medium has physical structure and allows bacteria to grow in physically informative or useful ways
(e.g. as colonies or in streaks). Solid medium is useful for isolating bacteria or for determining the colony
characteristics of the isolate.
2. Semisolid media
They are prepared with agar at concentrations of 0.5% or less. They have soft custard like consistency and
are useful for the cultivation of microaerophilic bacteria or for determination of bacterial motility.
3. Liquid (Broth) medium
These media contains specific amounts of nutrients but don’t have trace of gelling agents such as gelatin or
agar. Broth medium serves various purposes such as propagation of large number of organisms,
fermentation studies, and various other tests. e.g. sugar fermentation tests, MR-VR broth.
1. General purpose media/ Basic media:
Basal media are basically simple media that supports most non-fastidious bacteria. Peptone water, nutrient
broth and nutrient agar are considered as basal medium. These media are generally used for the primary
isolation of microorganisms.
Addition of extra nutrients in the form of blood, serum, egg yolk etc, to basal medium makes them enriched
media. Enriched media are used to grow nutritionally exacting (fastidious) bacteria. Blood agar, chocolate
agar, Loeffler’s serum slope etc. are few of the enriched media. Blood agar is prepared by adding 5-10% (by
volume) blood to a blood agar base. Chocolate agar is also known as heated blood agar or lysed blood agar.
3. Selective and enrichment media are designed to inhibit unwanted commensal or contaminating bacteria
and help to recover pathogen from a mixture of bacteria. While selective media are agar based, enrichment
media are liquid in consistency. Both these media serve the same purpose. Any agar media can be made
selective by addition of certain inhibitory agents that don’t affect the pathogen of interest. Various
approaches to make a medium selective include addition of antibiotics, dyes, chemicals, alteration of pH or a
combination of these.
Such media are called differential media or indicator media. Differential media allow the growth of more than
one microorganism of interest but with morphologically distinguishable colonies. Then, inoculations are
performed on medyum that have above properties. Inoculating mediums can be listed as follows.
The indicated bacterias are inoculated to some mediums in this experiment. Then, according to properties of
mediums, changes of mediums are observed. Also, changes of mediums give informations about properties of
bacterias. Thus, unknown bacterias are predicted.
RESULTS
For Experiment 1: Observing of Alpha hemolysis, Beta hemolysis and Gama hemolysis with Blood Agar
For Experiment 2: Inoculation Selective Media For Gram Positive Bacteria ( MSA or PEA)
For Experiment 3: Inoculation Selective Media For Gram Negative Bacteria ( MAC )
For Experiment 4: Inoculation Selective Media for Pathogenic Enteric Bacteria ( XLD Agar )
The identification of higher plants and animals involves the observations of the structural differences, both
internal and external, which exist among them. Most of these structural differences are visible to naked eye.
Even the identification of microscopic plants and animals involves the observations of their structural
differences under a microscope. Such identification based on structural differences is not possible in the case
of bacteria, because structural differences, which may differ from one species of bacteria to the other, are not
discernible even under a microscope. Because the structural differences in shape, size and arrangement are
only helpful in the process of identification, there are many species of bacteria having similar shape, size and
arrangement. Therefore, ultimately, the identification of bacteria is mostly based on the differences in their
biochemical activities. There are many biochemical tests for the identification of bacteria. We will talk about
the mechanics of these tests, their consequences, and the mistakes in their execution. The survival and
continued growth of microorganisms depend on an adequate supply of nutrients and a favorable growth
environment. For survival, most microbes must use soluble low-molecular-weight substances that are
frequently derived from the enzymatic degradation of complex nutrients. In this experiment, blood agar is
used to observing hemolysis in bacteria types. The mannitol salt agar is used to observing salt resistant in
bacteria types. The bacterias are choosed according to gram negative or positive and lactose positive or
negative properties in MCA, EA, SS and XLD. Only Pseudomonas species are choosen in Cetrimide agar and
Nalidixic acid. Some experiments are wrongly maked such as experiment 2. Staphylococcus epidermidis and
Staphylococcus aureus are gram positive bacterias. Although, E. coli and X bacteria are gram negative
bacterias, these bacterias have to colonies. We say that, experiment is contaminated. The reason of
contamination, inoculation loop is not cleared enoughly. So, some bacterias remain on the inoculation loop.
Thus, wrongly positive results are observed. The one of wrongly maked experiment is experiment 3. Mac
Conkey Agar suppresses the growth of Gram positive bacteria. But, S. aureus has colonies. The this situation
shows contamination of experiment. The reason of this contamination is lack of sterilization. The same
situation exists in experiment 5.
RESOURCES
https://www.alice.cnptia.embrapa.br/bitstream/doc/1055907/1/276891388061PB.pdf
https://www.easynotecards.com/notecard_set/80649
https://www.scribd.com/document/216995056/Cetrimide-Agar-Base