MICROBIOLOGY LAB

Modupe Epebinu 7

Identification And Analysis Of Klebsiella pneumoniae.

Modupe Epebinu

Baltimore City Community College

Professor Mark Dreyfuss

April 29, 2021

 

 

 

 

 

 

 

 

 

Identification And Analysis Of Klebsiella pneumoniae.

Introduction

Background information

Dealing with an unknown sample in the laboratory requires a number of tests for potential elimination processes. When given an unknown sample, one of the critical steps to take is ensuring that you have many portions of the sample. These enable one to carry out several tests where each test may infer the possibility of a number of pathogens or items under study. Therefore, examining the sample's nature helps to decide on the rightful tests to be carried out, whether in a solution or solid form. The number of tests determines the number of potions to be made; they should be enough for exhaustive confirmations and eliminations. The sample's nature plays a big role in whether you need to have test solutions or just a physical examination.

Purpose objective

Carrying out a test may give results that are not very specific. This is because some samples have similar characteristics. It may, therefore, not be ideal for making conclusions on findings that are depicting multiple inferences hence the need to have more tests to come up with a specific finding of the given sample. Similarities in samples' reactions can be very confusing if only a few tests are done for verification and to eliminate negative results and concentrate on positives that lead to the correct answer. The importance of identifying the organism is because of the need to understand the type of diseases or infections it causes and the potential remedies to the infections.

The materials and methods

In order to perform the experiment, the following materials were put at our disposal; API-20E analytical system: sterilized distal water, MacConkey agar culture, phosphate buffer saline, sterile Pasteur pipette, Caplets: CIT, VP, GEL, , ADH, ODC, IND, TDA, VP and LDC, sterile liquid paraffin , Kovac’s reagent, Ferric chloride and analytical profile index.

We performed the oxidase test before we inoculated.

We prepared a strip by doing the following; took 5ml of sterilized distal water and added to h holes of the tray so that the humid conditions required could be created, we placed the strip in the tray. We then prepared the bacterial suspension from the provided sample. We isolated four colonies from MacConkey agar culture. We suspended them in a test tube that we had filled with 5 ml of phosphate buffer saline.

We then inoculated the strip by doing the following; we filled the upper as well as the lower part of caplets with a bacterial suspension using a sterile pasture pipette of the following caplets; CIT, GEL, and VP. For all the other caplets, we filled only the lower part with the suspension to the concave line. We also filled the lower part with sterile liquid paraffin so that anaerobic conditions were provided to the caplets. These caplets include the, ADH, ODC, and LDC. We covered the band and incubated it for 24 hours maintaining.

After the period of incubation, we added the following reagents; we placed a drop of Kovac’s reagent to the IND caplet and made an immediate reading. We added a drop of Ferric chloride 10% to the TDA caplet and made an immediate reading. We added a drop of VP1 to VP caplet. We added a drop of VP2 and then read after 10 minutes.

We separated the test into 7 sets on the results page. We specified a number for each test as either 1, 2 or 4. We gave every positive result its number while assigning a 0 to a negative result. When we added the numbers correspondingly to the results within every group, we attained 7 digit profile numbers with which we compared them to the numbers found in the analytical index and documented the bacterium sample.

We observed all the characteristics and aspects of the procedure while recording the observations. This included the color of the reagents as well as the length of the growth samples in the agar plates.

Results and discussions

Test	Code	Negative result	Positive result
β-gala+C4:C23ctosidase	ONPG	Colorless	Yellow
Arginie Dihydrolysis	ADH	Yellow	Red-orange
Lysine Decarboxylase	LDC	Yellow	Red-orange
Ornithine Decarboxylase	ODC	Yellow	Red-orange
Citrate Utilization	CIT	Yellow	Red-orange
Hydrogen Sulfide	H2S	Colorless	Black sediment
Urease production	URE	Yellow	Red-orange
Tryptophan Deaminase	TDA	Yellow	Dark brown
Indole production	IND	Yellow ring	Red ring
Acetone production	VP	Colorless	Pink-red
Gel Hydrolysis	GEL	No pigments	Black pigments
Glucose	GLU	Blue	Yellow
Manitol	MAN	Blue	Yellow
Insonitol	INO	Blue	Yellow
Sorbitol	SOR	Blue	Yellow
Rhaminose	RHA	Blue	Yellow
Sucrose	SAC	Blue	Yellow
Melibiose	MEL	Blue	Yellow
Amayloid	AMY	Blue	Yellow
Arabinose	ARA	Blue	Yellow

To identify the isolates, large dome-shaped highly mucous colonies of the bacteria that fermented the lactose grow into a string measuring 6 mm in length. We prepared smears from the isolated colonies on a glass slide of which we then stained it using the Gram stain. Short rods that were gram-negative of the bacteria appeared in the stained smear. This confirmed that indeed the unknown sample was indeed Klebsiella pneumonia.

Klebsiella pneumonia does not affect healthy individuals. However, it badly affects people who have an immune system that is weak as a result of a medical condition of having been on long-term use of antibiotics (Vading, Nauclér, Kalin & Giske, 2018). The infections caused by these bacteria are treated by using antibiotics, however, some of the strains are highly drug-resistant. The bacteria is harmless while in the intestines of a human being. However, when they infect the body, the target area includes the lungs, the brain, eyes, bladder, blood, liver, as well as wounds. Each location results in a different type of symptoms as well as the treatment procedure. The infections include pneumonia, UTI, cellulitis, myositis, Meningitis, endophthalmitis, pyogenic liver abscess, and blood infection. 

Works Cited

Ceccarelli, G., Falcone, M., Giordano, A., Mezzatesta, M., Caio, C., Stefani, S., & Venditti, M. (2013). Successful Ertapenem-Doripenem Combination Treatment of Bacteremic Ventilator-Associated Pneumonia Due to Colistin-Resistant KPC-Producing Klebsiella pneumoniae. Antimicrobial Agents And Chemotherapy, 57(6), 2900-2901. DOI: 10.1128/aac.00188-13

Osman, E., El-Amin, N., Adrees, E., Al-Hassan, L., & Mukhtar, M. (2020). Comparing conventional, biochemical and genotypic methods for accurate identification of Klebsiella pneumoniae in Sudan. Access Microbiology, 2(3). doi: 10.1099/acmi.0.000096

Paterson, D., Mulazimoglu, L., Casellas, J., Ko, W., Goossens, H., & Von Gottberg, A. et al. (2000). Epidemiology of Ciprofloxacin Resistance and Its Relationship to Extended-Spectrum  -Lactamase Production in Klebsiella pneumoniae Isolates Causing Bacteremia. Clinical Infectious Diseases, 30(3), 473-478. DOI: 10.1086/313719

Rojas, L., Salim, M., Cober, E., Richter, S., Perez, F., & Salata, R. et al. (2016). Colistin Resistance in Carbapenem-Resistant Klebsiella pneumoniae: Laboratory Detection and Impact on Mortality. Clinical Infectious Diseases, ciw805. DOI: 10.1093/cid/ciw805

Vading, M., Nauclér, P., Kalin, M., & Giske, C. (2018). Invasive infection caused by Klebsiella pneumoniae is a disease affecting patients with high comorbidity and associated with high long-term mortality. PLOS ONE, 13(4), e0195258. doi: 10.1371/journal.pone.0195258

Yu, V., Hansen, D., Ko, W., Sagnimeni, A., Klugman, K., & von Gottberg, A. et al. (2007). Virulence Characteristics ofKlebsiellaand Clinical Manifestations of K. pneumoniae Bloodstream Infections. Emerging Infectious Diseases, 13(7), 986-993. DOI: 10.3201/eid1307.070187