Having the ability to collect and identify unknown microorganisms is vital in health and medicine. This capability is important for a variety of reasons, such as knowing the causative agent of disease, knowing if the microorganism obtains any beneficial properties and knowing the correct microorganism to use to create a successful antibiotic. Implementing the experimental methods learned thus far in the microbiology laboratory allowed an unknown bacterium to be identified as a result of this study.
This study allows the student to exercise their ability to use and understand sterile techniques, transmission, and prevention of microorganisms. Materials provided by the instructor, including an agar plate and cotton swab were used to collect this unknown. In this particular study the unknown bacterium was collected by swabbing the bottom of a tennis shoe belonging to the student, then immediately transferring the unknown to the agar plate by rubbing the swab across the surface.
Upon returning to the laboratory the growing bacterium was observed and then transferred to a new plate by using a three line streaking method illustrated by the professor. This streaking method was used to determine the purity of the unknown and to ensure that it was not a type of fungi. After completing the streak the plates were incubated and then observed to determine the form, elevation, and margin of the unknown. This observation resulted in a total of 20 colonies with the majority of them being yellow, circular, raised and entire. Following the observation of the incubated plates a Gram stain was performed.
The tools used to collect this data were provided by the professor and included: One bunson burner, one wire loop, one slide with slide cover, oil, and multiple staining chemicals. As directed by the professor the Gram stain was performed multiple times due to the fact that obtaining accurate results is extremely important. The results of each Gram stain performed indicated that this unknown bacterium was Gram positive with a coccus cell shape and staphlyo organization. Once the Gram stain was successfully completed with results ruling out pathogens the unknown bacterium underwent a series of biochemical tests.
This testing began with a catalase that was used to differentiate staphylococci from streptococci. This bacterium resulted in being catalase negative, which infers that the bacteria is streptococci that do not respire using oxygen as the final electron acceptor. This also concludes that the unknown bacteria may be anaerobes or facultative anaerobes that may only ferment. Another test performed indicated that the unknown bacterium was negative for arginine, meaning that the bacterium cannot use the enzyme arginine as a source of carbon and energy for growth.
The triple sugar test, simply known as TSI, was also performed and indicated that the bacterium was fermenting and producing organic acids, which caused the color of the top and bottom of the slant to be yellow. Given that the entire slant remained yellow it is assumed that the bacteria is fermenting Glucose and Lactose or Glucose and Sucrose. Another test that was performed on the bacterium multiple times due to inactivity was the Litmus Milk test. This test had to be observed twice on different days, to ensure accurate results.
Both Litmus test observations resulted in a neutral bacterium that does not digest casein, make yogurt, release alkaline products, or release organic acids. The biochemical tests continued throughout the semester with two other tests one called MIO and the other is known as SIM. MIO is a test used to assess the motility, indole production, and ornithine-decarboxylase activity. While the SIM test is used to assess the sulfide production, indole formation, and motility. Both tests indicated that the bacterium is a non-motile facultative anaerobe that does not react with Indole and is negative for both eating ornithine and sulfide production.
The last two tests that will be discussed seemed to play a significant role in identifying the unknown bacterium using Bergy’s Flow Chart. The first of these tests is the blood agar test that showed the bacterium was negative for eating heme’s, which lead to observing the bile esculin test that is used to differentiate enterococci and group D streptococci from non group D viridans group streptococci. The results of the bile esculin test revealed that the unknown bacterium is positive and has the ability to breakdown esculin, which allows it to filter out and reduce the amount of bacteria.
In conclusion, after completing many various differential tests and reviewing the Bergy’s Flow Chart the unknown bacterium has been determined to be gram-positive Enterococcus faecium. E. faecium is a prokaryote that normally grows in groups or chains and is considered to be vancomycin resistant. Although, E. faecium is typically found in the gut of humans and animals it can be an opportunistic pathogen and may cause infection when the immune system has been damaged.
Due to their ability to resist a variety of antibiotics and survive on inanimate objects for long periods of time, they have recently been termed an important nosocomial pathogen referred to as a “superbug”. Infections related to E. faecium include urinary tract infections, wound infections, and soft tissue infections. In a patient who has previously damaged cardiac valves E. faecium may lead to endocarditis. Although Enterococci are part of the normal flora in humans and animals there are only a few of its species that cause clinical infections.
The most common being Enterococcus faecalis, which accounts for up to 90% of clinical isolates. However the other species, E. faecium, particularly the vancomycin-resistant strain that was once nearly 10%, is beginning to increase in the amount of clinical isolates. This increase may be a result of surgical wounds, intravascular catheter placement, contaminated food, and poorly treated sewage. Considering the fact that E. faecium may be excreted in human, pets, and wild animal feces it is to be expected that this bacteria would be found on the bottom of any ones tennis shoes.
Since this type of bacteria has the ability to live on inanimate objects for long periods of time it is important to take precautions to prevent illness due to contact with this bacteria. Preventions are as simple as maintaining basic hygiene, such as effective hand washing after using the restroom or while handling any type of feces. It is extremely important for healthcare workers to prevent the spread of E. faecium by continuously cleansing their hands thoroughly, since they are often dealing with many different patients with different illnesses, wounds, and procedures.
