PSEUDOMONAS Unknown Bacteria

DECEMBER 11, 2014 UOIT

Vita Koren

MORPHOLOGY CHARACTERIZING

Vita Koren 12-11-2014

Acetobacter Acinetobacter

Alcaligenes Bacillus

Citrobacter Clostridium Enterobacter

Klebsiella Lactobacillus

Micrococcus Pediococcus Pseudomonas

Staphylococcus Trichococcus

Acetobacter Acinetobacter

Alcaligenes Citrobacter

Enterobacter Klebsiella

Pseudomonas

Bacillus Clostridium

Lactobacillus Micrococcus

Pediococcus Staphylococcus Trichococcus

Gram Stain - +

Acetobacter

Acinetobacter

Alcaligenes Pseudomonas

Growth under Aerobic/ Anaerobic

+ for Aerobic Faculative Anaerobe Anaerobic -

Acetobacter Alcaligenes

Pseudomonas

Acinetobacter

Enterobacter Citrobacter

Klebsiella

Motility

+ Motile - Non-motile

BIOCHEMICAL TESTS

Acetobacter

Pseudomonas

Alcaligenes

Oxidase test

- (No color) + purple

Pseudomonas

Glucose OF Basal

+ for oxidation - for oxidation and fermentation

Alcaligenes

SUMMARY OF RESULTS

Test Performed Result Obtained Interpretation of Result

Plate Characterization - Colonies are circular - Convex - Entire - No distinct color - Solid colored, not clear - Small colonies

Morphological Features observed with the naked eye

Negative Staining - 3.0 um - Rod shaped - Fusiform

Bacteria is bigger than other bacteria

Simple Staining - 2.5 um - Rod shaped - Fusiform

Confirm that size is of larger value than other bacteria, the shape and the arrangement

Wet Mount - Rod shape - Darting around

Further confirm shape Motile

Culture Method (LB Broth) - Pink on surface of tube - Grew away from stab line

Motile Probably Aerobic

Growth under aerobic/anaerobic Growth at the top Strict Aerobe Gram Stain Pink Confirmed gram negative bacteria

Oxidase Test Dark blue color change Oxidase Positive

Glucose OF Basal Test - Yellow color change on surface of tube with no oil - No color change in tube with oil

Oxidation of glucose positive Fermentation of glucose negative

Catalase Test Bubbles Catalase Positive Endospore Test Red No endospore formation

Before performing any tests, the bacteria was observed on the plate to have circular colonies, a convex elevation, no distinct color and the colonies were small. The microscopic tests

performed were the negative stain and the simple stain to determine size, shape and arrangement of bacteria. The bacteria were rod shaped and in a fusiform arrangement. As well, the bacteria

were on the larger side (2.5-3 um). Following the first flowchart, the gram stain resulting in a pink stain resulted in the bacteria being gram negative, eliminating gram positive bacteria. Next, growth under aerobic, faculatively anaerobic and anaerobic conditions demonstrated which

bacteria were strict aerobes compared to those that were facultative anaerobes. My bacteria were aerobic because they only showed growth at the surface. At this point, only Acetobacter,

Acinetobacter, Alcaligenes and Pseudomonas were left as viable options to be my gram negative, strict aerobic bacteria. Finally, motility was able to leave the options as Acetobacter, Alcaligenes and Pseudomonas. Motility was tested with the culture method that resulted in the bacteria

moving away from the stab line. The wet mount demonstrated the bacteria had motility which was characteristic of these 3 bacteria. Next, the oxidase biochemical test was then performed and

a color change to purple indicated a positive result eliminating Acetobacter which would remain clear due to no cytochrome oxidation. Alcaligenes and Pseudomonas test positive for the oxidase test. To conclude, a glucose OF basal test was able to differentiate between Alcaligenes and

Pseudomonas because Alcaligenes cannot oxidize or ferment glucose whereas Pseudomonas is able to oxidize glucose as indicated by a color change to yellow at the surface of the test tube

without oil. With that, the unknown was identified as Pseudomonas! Additionally, Pseudomonas was further confirmed with a positive catalase test and a negative endospore forming test.

To further prove the determination of Pseudomonas, the citrate test can be performed and a

positive result would be obtained due to the green solution turning blue (Palleroni, 2010).

Additionally, a blood agar test can be performed and there would be beta-hemolysis of the agar

due to Pseudomonas whereas Alcaligenes would not lyse it. (Buxton, 2010).

References

Buxton, R. (2010). “Blood Agar Plates and Hemolysis: Non-Fermenting Gram-Negative Rods”

American Society for Microbiology 11 (2): 899-905

Palleroni, N. J. (2010). "The Pseudomonas Story". Environmental Microbiology 12 (6): 1377–

1383.