Bacterial Growth and Cultivation
Bacterial growth, also referred to as cultivation, takes place in a laboratory. These bacteria are raised in or on many types of microbiological media. Regarding the kind of bacteria it is used to isolate or identify, each medium has a different and special combination of nutrients. NaCl, carbohydrates, proteins, growth factors, pH buffers that balance the medium's pH, indicators that show pH changes, and indicators of pH changes are just a few of the nutrients that can be added to broths. The cookbook approach was the traditional method of creating media. The main objective of this lab is to run selective and differential tests on Escherichia coli, Enterococcus durans, pseudomonas fluorescens, staphylococcus saprophyticus, and Enterobacter aerogenes in order to establish such characteristics like the ability to ferment carbohydrates and production of gases. The knowledge of Culturing is very important to clinicians like us because it will help us to isolate and identify the type of bacteria strain and also its Abundance in a sample, this is very important in primary diagnosis and determination of an infectious disease. (Thagard, 2000, pg 161)
Materials
Broth culture of each of the following bacteria: 1. Escherichia coli 2. Enterococcus durans, 3. Pseudomonas fluorescens 4.Staphylococcus saprophyticus 5. Enterobacter aerogenes (Gram negative rod) 1 – Mannitol Salt Agar (MSA) plate 1 – Hektoen Agar plate 1 – Eosin Methylene Blue agar plate 2 - SIM agar tubes 2 – Bile Esculin 2 – Phenol red glucose tube 2 – Phenol red lactose tube 1 – Bottle of hydrogen peroxide 1 – glass slide Striker Bunsen burner Wire inoculating loop.
Procedure
Mannitol Salt Agar
( Engelkirk, Duben-Engelkirk, 2008, Pg 189)
Organism 1 2 3 and 4 were used
Dividing the MSA plate into four equal sections using a marker pen
Labelling of each section with the organisms name, our names and lab time.
We them inoculated each section with an organism using a sterilized loop gently.
Inversion of the plate and incubation at 37 degrees until next lecture
Recording of results
Eosin Methylated Blue Agar
(Dworkin and Falkow, 2006, Pg 161)
Organisms 1, 3, 4, and 5 were used
Dividing of EMB plate using a marker pen into four sections
Labelling of the sections with organism name, group name, and lab time.
Inoculation using a sterilized loop
Incubation at 37 degrees until next lecture.
Observation and recording
Hektoen
Organisms 1, 3, 4, and 5 were used
Dividing of EMB plate using a marker pen into four sections
Labelling of the sections with organism name, group name, and lab time.
Inoculation using a sterilized loop
Incubation at 37 degrees until next lecture.
Observation and recording.
SIM Tube
Organisms 1 and 4 were used
Labelling of SIM tubes with organism name, group name, and lab time.
Inoculation of media using an inoculation needle
Incubation at 37 degrees
Observation and recording of results
Catalase Test
Organisms 1, 2, 4, and 5 were used
Draw four circles on a glass microscope slide
Place a loop of bacteria in each and add water
Water bubbles indicate possible reaction
Observe and record.
Bile Esculin
Organisms 1 and 4 were used
1. Labelling of Esculin tubes with group name, organism name, and lab time
2. Inoculation using a sterile loop.
3. Incubation at room temperature
Conclusion
In Manitol salt agar staphylococcus saprophyticcus ferments Mannitol turning the medium yellow and is selected other bacteria didn’t grow. The medium is selective thus the staphylococcus can tolerate high salt conditions. In the EMB agar staphylococcus saprophyticus does not grow because it doesn’t ferment lactose, positive and negative lactose colonies are differentiated thus it is both selective and differential. In the Hektoen experiment staphylococcus saprophyticus does not show growth because it is gram positive, yellow colonies are observed for other bacteria because of their fermenting power. It is a selective media. In SIM experiment Escherichia coli appeared dark in color along the line of inoculation, motility, and red colonies were also observed because the organisms produced sulphide. It is a differential agar. In catalase test exp, staphylococcus saprophyticus showed effervescence due to production of oxygen. It is a selective agar. The bacteria use oxygen to respire. In Bile Esculin experiment Escherichia cocci medium changes its color to black, the bacteria hydrolyzes esculin. In this experiment it is used selectively. In the Fermentation tube experiment Escherichia coli changes the color of the medium and produces a gas. The bacteria have the ability to ferment carbohydrates. The medium is selective.
References
Thagard, P. (2000). How scientists explain disease. Princeton, NJ [u.a.: Princeton Univ. Press.
Engelkirk, P. G., & Duben-Engelkirk, J. L. (2008). Laboratory diagnosis of infectious diseases: Essentials of diagnostic microbiology. Baltimore: Wolters Kluwer Health/Lippincott Williams & Wilkins.
Dworkin, M. M., & Falkow, S. (2006). Proteobacteria: Gamma subclass. New York, NY: Springer.
