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ABOUT THIS MICROBE
ABOUT THIS MICROBE
Klebsiella pneumoniae was first documented in 1882 by German physician and bacteriologist, Carl Friedlander. He isolated the bacteria from the lungs of patients that had recently passed away from pneumonia and described the microorganism as an encapsulated bacillus. [1]
Klebsiella pneumoniae has a biosafety level of 2, meaning it poses moderate hazards for laboratories and the environment due to its pathogenic nature.
To grow K. pneumoniae, you must use Trypticase Soy Agar specifically labeled medium #18 and incubate it in an aerobic environment.
TAXONOMY
TAXONOMY
Taxonomy of K. pneumoniae [2]
Kingdom: Bacteria
Subkingdom: Negibacteria
Phylum: Proteobacteria
Class: Gammaproteobacteria
Order: Enterobacteriales
Family: Enterobacteriaceae
Genus: Klebsiella
Species: Klebsiella pneumoniae
VISUALIZING THE MICROBE
VISUALIZING THE MICROBE
Klebsiella pneumoniae is a gram-negative bacterium that appears pink under a microscope due to its thin peptidoglycan layer which denotes its infectious qualities and tendency to be resistant to drugs. [3]
Klebsiella pneumoniae is rod-shaped, thus it belongs to the Bacillus group.
SEM - Scanning Electron Microscope
A Scanning Electron Microscope (SEM) is used by scanning the surface of a specimen with a focused beam of electrons, generating a variety of signals that contain information about the composition and topography of the sample itself.
This given sample was fixed, dehydrated, and then coated with a 1:1 mixture of gold and palladium. It was then viewed under a Sigma VP40 field emission SEM [4].
TEM - Transmission electron microscope
The use of a Transmission Electron Microscope (TEM) involves the technique of transmitting a beam of electrons through a specimen in order to conjure up an image.
The colony biofilms were fixed with 5% glutaraldehyde and then stained with osmium tetroxide (1%). After this primary stain and being washed with water, the specimen was stained with 1% uranyl acetate-1% phosphotungstic acid [5].
CLSM - Confocal Laser Scanning Microscope
The use of a Confocal Laser Scanning Microscope involves taking images of specimen from the focal plane, point by point with the use of laser excitation.
The specimen was stained with the Live/Dead BacLight Reagent where the live cells can be determined by their green hue while the dead cells are tinted red [6].
Gram Stain
SEM
TEM
CLSM
DIAGNOSTIC TESTS
DIAGNOSTIC TESTS
Blood Agar
Klebsiella pneumoniae does not produce exotoxins that destroy red blood cells. It participates in Gamma Hemolysis which is evident in a plate which has little to no marked change on its surface [7].
MacConkey Plate
Klebsiella pneumoniae’s growth on a MacConkey Plate indicates that it is gram-negative and that it has the ability to ferment lactose (positive). On a MacConkey Plate, this microbe produces pink colonies because of the lactose fermentation and subsequent creation of a acidic environment with a pH of less than 6.8 [9].
Mannitol
Mannitol Salt Agar Plates contain 7.5% NaCl concentration, thus Klebsiella pneumoniae who is positive for this test is halophilic and tolerant to environments with high salt concentrations. The yellow hue on the plate denotes mannitol fermentation [10].
DNase
The DNase plate is used to determine a bacteria’s ability to hydrolyze DNA. An organism that is found negative in the DNase test, like Klebsiella pneumoniae, will be evident with a plate that is green throughout and an absence of colorless zones around the colonies that are formed [11].
Anaerobic Chamber
Klebsiella pneumoniae is a facultative organism, meaning it can survive in aerobic conditions through respiration and can also survive anaerobic conditions through fermentation. Facultative organisms, seen in Tube #2, show great concentration of growth at the top of the medium when in the presence of oxygen [12].
Phenol Red Broth
Klebsiella pneumoniae is found positive for the Phenol Red Broth Test. Due to its ability to create acidic environments, it turns the phenol broth from red to yellow. K. pneumoniae also ferments glucose and thus will produce gas, as shown in Tube A [13] .
SIM Medium Deep Agar
Klebsiella pneumoniae is negative for the Indole exam, this is evident because it does not turn the solution red when Kovac Reagent is added. Being negative means that this microbe does not decompose the amino acid tryptophane into indole [14].
Klebsiella pneumoniae is negative for the Sulfur Reduction Test, known because it does not turn the solution black. This lets us know that H2S gas does not combine with the iron in ferrous ammonia sulfate in the presence of the microbe [16].
Klebsiella pneumoniae is negative for the Motility Test, thus it does not contain flagella or pilli that help it move. A lack of cloudiness in the tube with the microbe and medium denotes immobility [18].
Catalase
Bacteria that produce the enzyme catalase help them in facilitating cellular detoxification and repairing oxidative damage. Klebsiella pneumoniae is found positive for this test, as a result of its immediate effervescence (bubbling) in the presence of hydrogen peroxide [19].
Molecular Diagnostics
In order to run a PCR, both a forward and reverse primer is necessary.
The forward primer used was K16SF, a 15-mer found from positions 44 to 58 bp with following sequence: 5’- AGCACAGAGAGCTTG – 3’. The Melting Point of the forward primer was 41.9 °C and its GC content is 53 % [20].
The reverse primer used was K16SR, a 16-mer found from positions 155 to 170 bp with following sequence: 5’ – TGGGGAGTACGGCCGC – 3’. The Melting Point of the reverse primer was 53.6 °C and its GC content is 75% [20].
MacConkey Plate [8]
Mannitol [10]
DNASE [11]
Anaerobic Chamber [12]
Phenol Red Broth [13]
Sulfur Reduction Test [15]
Motility Test [17]
Catalase [19]
DISEASE AND TREATMENTS
DISEASE AND TREATMENTS
Klebsiella pneumoniae is an opportunistic pathogen that causes a plethora of diseases and infections including: urinary tract infection (UTI), pneumonia, bloodstream infection (BSI), meningitis, pyogenic liver abscess (PLA), etc… [21]
Klebsiella pneumoniae has been found to have mortality rates as high as 50% and is specially harmful for geriatric patients and those who are immunocompromised. [22]
It is traditionally found in healthcare settings and is today considered the most common cause of hospital-acquired pneumonia in the United States [1].
K. pneumoniae is primarily spread from person-to-person contact. Therefore, the CDC stresses the importance of strict adherence to proper hand-washing and the use of PPE when treating patients. Proper hand-washing protects patients and prevents health care workers from transmitting it through infected catheters, needles, or ventilators. [23]
According to the Gideon database, Abacavir is an antiviral drug used to treat HIV that has the potential to work against Klebsiella pneumoniae.
However, it is important to note that K. pneumoniae is becoming exceedingly resistant to antibiotics as the European Centre for Disease Prevention found that “more than a third of the K. pneumoniae isolates were resistant to at least one antimicrobial group.” [24]
ABOUT THE RESEARCHER
ABOUT THE RESEARCHER
Angelica Ruiz is a second-year Microbiology and Cell Sciences major from Miami, Florida who is minoring in Nutritional Sciences and Business Administration. She aspires to attend medical school and become a dermatologist and own her own clinic in the distant future. Her favorite hobbies include long-walks to the fridge and curling up on the corner of the couch as she wanders into the fantasy worlds where her novels are set in. She is a proud Cuban-American who loves introducing people to her culture, especially the music and cuisine from her gorgeous Caribbean Island.
REFERENCES
REFERENCES
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