There are legion grounds for placing unknown bacteriums. Some of these beings have distinguishable qualities that set them apart from one another. such as the exposure to certain environments. Through out the semester in the research lab. we are able to meet some of the few micro-organisms that we as worlds have come into contact with. With the cognition gained from the Sessionss in the research lab. we can now incorporate what we have learned to the procedure of happening out the unknowns given.
Materials and Methods
The professor gave out the unknown specimens. It contained one-?gram positive and one-?gram negative bacteriums from the given list. I was assigned unknown A. The procedure of designation was achieved by using processs learnt during the semester. Procedures were followed as stated in the lab manual ( 1 ) . Since the sample contained two unidentified bacteriums. the first measure was to insulate each bacteria utilizing streak home base technique. Tryptic Soy Agar ( TSA ) home base. and differential media such as Osmitrol salt and Eosin methylene blue ( EMB ) were used for isolation run technique.
This measure is imperative because the bacteriums need to be separated and isolated before they can be identified. Furthermore. gm staining was used to understand the basic morphology of these bacteriums. As these home bases were incubated and grown. the presence of two separate bacteriums settlements was seeable. The settlements from the Osmitrol salt were used to incubate a TSB stock to turn the gram-?positive civilization. The pureness of this stock was tested utilizing gram-?staining technique. A round settlement from the TSA home base was used to incubate a TSB stock for gram-?negative growing. Similarly. analyzing the morphology of the bacteria-?using gm staining technique tested the pureness of the both.
After the isolation of gram-?positive and gram-?negative bacteriums from unknown A. specific biochemical trials were performed. The consequences of the biochemical trials along with deductive logical thinking and riddance led to the designation the unknown bacteriums. The undermentioned trials were performed on the gram-?positive bacteriums: 1. Mannitol salt streaking 2. Gram stating of the pure isolate 3. Oxidase trial 4. Catalase trial 5. Coagulase test The undermentioned trials were performed on the gram-?negative bacteriums: 1. Gram staining of the pore isolate 2. Blood agar home base streaking 3. SIM ( Sulfide. Motility. Indole ) trial 4. Catalase trial Consequences
Table 1. Biochemical Trials for the Gram-?Positive Unknown TESTS PURPOSE REAGENTS/MEDIA OBSERVATION RESULTS Gram discoloration To find the gram reaction and morphology ?Crystal violet. Iodine. Alcohol. Safranine
Purple coccus. connected Gram positive purple coccus
Mannitol salt Selective growing media for Staphylococci and Micrococcaceae
Mannitol salt home base and gm positive isolate stock After incubation. media turned xanthous
Bacteria is mannitol salt positive
Oxidase trial To find if bacterium produces cytochrome degree Celsius oxidase
No colour alteration Bacterium is oxidase negative Catalase trial To place if bacteria produces catalase
Hydrogen peroxide Bubbling is seen Bacterium is catalase positive Coagulase trial Used to place if bacteria produces coagulase ( enzyme that coagulums blood plasma )
Citrated coney plasma Clouding and hardening of plasma is seen Bacterium is coagulase positive
Table 2. Biochemical Trials for the Gram-?Negative Unknown TESTS PURPOSE REAGENTS/MEDIA OBSERVATION RESULTS Gram discoloration To find the gram reaction and morphology
Crystal violet. Iodine. Alcohol. Safranine
Small pink rods Gram negative pink rods Blood agar plateSelective growing media
Blood agar home base After incubation. media displayed beta haemolysis. metallic shininess. and blue-?green pigment growing ( fig 1 )
Non fermenting gram negative rods Oxidase trial To find if bacterium produces cytochrome degree Celsius oxidase
Oxidase reagent ( tetramethyl-?p-?phenyldiamine )
Color alteration to dark bluish Bacterium is oxidase positive
Catalase trial To place if bacteria produces catalase Hydrogen peroxide Bubbling is seen Bacterium is catalase positive
SIM ( Sulfide. Motility. Indole ) trial 1. To find the ability of an being to emancipate H sulphide ( H2S ) from
SIM medium and indole reagent The medium showed no colour alteration. motility. or colour alteration when indole trial was done.
-?/-?/-? The bacteria is sulfide. motility. and indole negative.
sulphurbearing aminic acids bring forthing a seeable. black color reaction. 2. To find the ability of an being to divide indole from the tryptophan molecule. 3. To find if the being is motile or non-?motile. Discussion and Decisions:
The biochemical trials performed on the gram-?positive bacteria worked consistently to contract down the possible species. and finally extinguish every being on the list except the right 1. The gm discoloration. that showed the gm positive coccus and the transmutation of the mannitol media to yellow colour left me with two picks that fit this profile-? Micrococcus luteus and Staphylococcus aureus. Biochemical trials that differentiate these two were performed. These included. oxidase. catalase. and coagulase trial. The consequences of
these trials ( oxidase negative. catalase positive. coagulase positive ) . confirmed that the gram-?positive micro-organism in unknown A is Staphylococcus aureus. After the Gram+ bacteria was identified as such. farther trials were performed to contract down which peculiar Gram- species this was. The first trial done was blood agar home base. The consequences show that after incubation the blood agar media displayed beta haemolysis. metallic shininess. and blue-?green pigmentation.
Colonies of Pseudonomas aeruginosa frequently display similar consequence on this medium ( 2 ) . To corroborate the illation. oxidase. SIM. and catalase trials were performed on the gram-?negative isolate. The consequences showed that the bacteria was oxidase positive. SIM negative ( -?/-?/-? ) . and catalase positive. Similar consequences are characteristic of P. aeruginosa. Hence. the gram-?negative micro-organism in unknown A is Pseudonomas aeruginosa. Appendix:
Fig. 1. Photograph of the blood agar medium ( after incubation ) streaked with the gram-?positive isolate of unknown A. References ( 1 ) Harley. J. P. ( 2014 ) . Lab Exercises in Microbiology. University of Kentucky: McGraw Hill
( 2 ) Buxton. R. ( 2010 ) . Blood Agar Plates and Hemolysis: Non-?Fermenting Gram-? Negative Rods ( including Pseudomonas aeruginosa ) .
Retrieved from hypertext transfer protocol: //www. microbelibrary. org/library/laboratory-?test/2862-?blood-?agar-?plates-?and-?hemolysis-?non-?fermenting-?gram-?negative-?rods-?including-?pseudomonas-?aeruginosa ( 3 ) Summary of Biochemical Tests. ( n. d. ) . Retrieved from hypertext transfer protocol: //www. uwyo. edu/molb2210_lab/info/biochemical_tests. htm ( 4 ) Sigma Aldrich. J. S. ( n. d. ) . Pseudomonas Media and Tests. Retrieved from hypertext transfer protocol: //www. sigmaaldrich. com/technical-?documents/articles/analytix/pseudomonas-?media. hypertext markup language
