Shannon Carr

Microbiology for Nurses

May 6, 2014
Lab Report 2- Unknowns
Abstract
This lab was done to discover our unknown bacteria. I received the
unknown bacteria, unknown 12. I performed a series of tests to
discover what bacteria unknown 12 was. I performed a gram stain after
streaking for single colonies on unknown 12. Then after finding that it
was gram negative and a bacilli shaped bacteria, I performed an
oxidase test. The oxidase test resulted positive, so then I performed a
glucose test. The glucose test was positive, so my unknown bacterium
was vibrio cholerae.

Introduction
The purpose of this lab is to perform several different tests and know
how to do them in order to discover your unknown. According to
Steven Chu of Science AAAS, Vibrio cholerae biofilms displayed three
distinct levels of spatial organization: cells, clusters of cells, and
collections of clusters, (Chu 2012). When streaking for single colonies I
chose a larger, whiter colony as opposed to the yellow smaller
colonies. It was important to get a perfect single colony, so that the
tests are accurate enough to find he right bacteria. Being a nurse, it is
important to know what each bacterium is, and what it causes. That

way the nurse will be able to prescribe the right antibiotic and the right
amount. According to The New England Journal of Medicine, Cholera is
a diarrheal disease caused by colonization of the intestines by cholera
toxinexpressing strains of the waterborne enteric bacte- rium V.
cholerae. An outbreak can arise suddenly, especially in vulnerable
populations with compromised sanitation infrastructure, (Devault
2014). Knowing how to do these tests and identify what they mean is
important when discovering a bacteria. According to microbiologist
Mike Son, Vibrio Cholerae is spread in Haiti by poor infrastructure, poor
sanitation, and bad human practices because there is no education in
Haiti about Cholera. If they had the right equipment in Haiti to know
what causes Vibrio Cholerae and how to determine if it is cholera, they
would not have this problem. This is exactly why we did the tests in lab
when determining our unknown. If we didnt do this kind of testing in
America, we would have the same problem. The purpose of this lab
was to be able to preform these tests to find out unknown bacteria.

Methods and Materials

In Lab 2, I performed several tests to figure out my unknown bacteria
following the dichotomous key. After streaking for single colonies, I did
a gram stain of one of the larger, whiter colonies. The gram stain was
negative, so I then checked the shape of the bacteria and it was bacilli.
Then I performed an oxidase test and it resulted in positive. Next, I

performed a glucose test, which was also positive, telling me that my

unknown bacterium was Vibrio Cholerae, (Son 2014).

Results
In Lab 2, after streaking for single colonies of my unknown bacteria, I
saw that there were two types of colonies: larger, white, round ones;
and smaller, yellow, round ones. I took a larger white one when doing
the gram stain. The gram stain came out red making the bacteria gram
negative. The individual particles were bacilli shaped. The next test
was the oxidase test, which resulted positive due to its purple coloring.
The last test was the glucose test, which was also positive because it
was yellow. Table 1 shows each test performed, its results, and the lab
used. Table 2 gives the colors that told me the results.

Table 1
Test
Gram Stain
Oxidase
Glucose

Results
Negative
Positive
Positive

Table 2
Test
Gram stain
Oxidase
Glucose
Figure 1
Vibrio Cholerae Gram Stain

Lab Number
Lab 5
Lab 8
Lab 7
Color of Result
Red
Purple
Yellow

(Example of a gram-negative stain with bacilli shaped bacteria)

Discussion and Conclusion
After completing Lab 2, The Unknowns lab, I have come to the
conclusion that my unknown bacterium is Vibrio Cholerae. To figure out
that my unknown bacteria was Vibrio Cholerae, I had to perform three
tests. Before doing any of the tests, I had to streak for single colonies
and pick a colony that would show accurate results. I picked one of the
larger, whiter colonies, as opposed to a smaller, yellow. The colony had
a circular form, convex elevation, and entire margin. After streaking for
singles colonies, I did a gram stain. Examining under the microscope, I
saw that my bacteria was red and bacilli shaped. This told me that I
had a gram negative bacteria. Gram Negative bacteria have an outer
membrane, which contains Lipopolysaccharides; therefore, I knew my
bacteria contained Lipopolysaccharides, (Son, 2014). After I knew that
my bacterium was gram-negative, I performed an Oxidase test.
According to Lab 8 Biochemical Assays, the oxidase test detects the
presence of cytochrome oxidase through the use of the artificial

electron acceptor N,N,N,N-tetramethyl-p-phenylenediamine. This

compound changes colors from yellow to purple when it accepts the
electrons from the cytochrome c, which gets reduced by the
cytochrome oxidase. As seen in Table 2, my swab turned purple, so I
knew that my oxidase test was also positive. Next, following my
dichotomous key, I performed a glucose test. The glucose is done to
determine whether or not there is glucose present in the bacteria. My
bacteria did have glucose in it. I know this because my tube was
yellow, which means positive. My results did not show any gas
(hydrogen or carbon dioxide) in the bacteria, which can result in the
presence of glucose, (Son, 2014). Vibrio Cholerae in many strains can
cause Cholera. According to AAAS Science, Large increases in the
production of cerebrospinal fluid have been observed after the
intraventricular administration of cholera toxin. Because cholera toxin
stimulates adenylate cyclase, the data suggest that adenosine 3',5'monophosphate plays a role in cerebrospinal fluid production, (MH
Epstein, AM Feldman, SW Brusilow, 1977). Because Cholera can be
lethal and spread, it is important for microbiologists to be able to
identify when it is present. The overall purpose of this lab was to
determine our unknown bacteria from performing tests, to prove that
we learned out to do them in lab.
References
Colwell, R., Kaper, J., & Joseph, S. (1977). Vibrio cholerae, Vibrio
parahaemolyticus, and Other Vibrios: Occurrence and Distribution

in Chesapeake Bay. AAAS Science, 198, 4. Copy & Paste |

Parenthetical
Devault, A., & Golding, B. (2014). Second-pandemic strain of Vibrio
cholerae from the Philadelphia cholera outbreak of 1849. The New
England Journal of Medicine, 1, 10. Copy & Paste | Parenthetical
Epstein, M., & Feldman, A. (1977). Cerebrospinal Fluid ProductionL
Stimulation by Cholera Toxin. Science AAAS, 196, 2.
Google Images. (n.d.). Google.com. Retrieved May 5, 2014, from
http://2.bp.blogspot.com/tJde0RfK2eE/USNzJ2uquLI/AAAAAAAAAC0/
Son, M.S. 2013. Microbiology Lab Manual. 2nd Edition. Academx
Publishing.