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Microbiology Concept Inventory
Please note, you must be an educator in higher ed or maybe high school to qualify to recieve the MCI
( 49351 Reads)
Bacteriophages are specific in action. A particular phage may be very specific in that it will infect only a few strains of a certain bacterial species. On the other extreme, another phage may infect strains of two or more species of a particular genus. Susceptibility to lysis by a particular phage may be the only apparent phenotypic difference between two bacterial strains and may be the only means by which a strain causing an outbreak of disease can be recognized. This observation is the basis for phage-typing, a procedure for characterizing and detecting bacterial strains by their reaction (susceptibility or resistance) to various known strains of phages. The phage-typing procedure is sometimes employed in epidemiology to identify and trace the progress of an infectious agent This has been a standard procedure for strains of Staphylococcus aureus involved in outbreaks of food poisoning or other infections. A species-specific phage is used routinely in the identification of Bacillus anthracis, the causative agent of anthrax. Thus, reproduction of anthrax in laboratory animals is not necessary to confirm identification of this organism.
Period 1
Materials
Broth cultures of eight potential host strains including several strains of Escherichia coli
Suspensions of phage strain JL-1 and another phage strain (each appropriately diluted)
4 plates of Bottom Agar (well-dried)
Micropipette (P20 preferred) and sterile tips
8 sterile swabs
- Label one half of each plate for one of the 8 bacterial cultures provided. For each culture, make a single, broad (at least 1 inch wide) streak with a cotton swab on the appropriately-labeled half of the dish; be sure to keep the streaks separate. Discard each swab into the disinfectant after streaking.
- Once the streaks have dried completely (this may take some time), spot-inoculate 10 µl(with the micropipette) phage JL-1 on each streak, near one end - dropping just a tiny amount of the phage suspension from the pipette tip. If you touch a streak, be sure to replace the tip, as you do not want to cross-contaminate the cultures.
- Repeat the above procedure with a fresh pipette tip and the other phage - spot-inoculating this phage near the other end of each streak.
- Allow the phage inocula to dry completely. Incubate at 37°C for 1-2 days (or 30°C if incubation is more than 2 days).
Period 2
Figure 9.3. Phage-typing. The first and second plates from the phage-typing experiment. Test strains were Esherichia coli A, Esherichia coli B, Esherichia coli K, and an orange mutant of Esherichia coli. The two phage were JL-1 and KX-32. Interpret the results and record your observations.
Figure 9.4. Phage-typing 2. The third and fourth plates from the phage-typing experiment. Test strains were Klebsiells oxytoca, Klebsiella planticola, Esherichia coli F-, and Esherichia coli Hfr. The two phage were JL-1 and KX-32. Interpret the results and record your data.
- Examine the plates for evidence of lysis (a giant plaque) in the area where a phage was inoculated. Tabulate results below, recording + for lysis (=sensitivity of a bacterial strain to a particular phage).
- From our limited results, which phage appears to be species and strain-specific? Which appears to be just species-specific? To answer these questions with certainty, we would have to test dozens or hundreds of strains of each species and related species! (Remember that if you do a similar experiment out in the real world!)