Lactobacillus, using the spread plate method. MRS agar plates are
incubated at 37°C for 48 hours. Cultures can then be stored at
4°C until the next lab session. This can be done in about one
Students are asked to observe their plates and are reminded to
use the one that has between 20 and 200 colonies. The other
plates are called TFTC (too few to count) or TNTC (too numerous to count). See Discussion for examples of results. Students
then calculate the CFUs per fly by multiplying the number of colonies on the plate by the reciprocal of the dilution factor (Keler,
2010). For example, 50 colonies on the 10− 4 plate would be
50 × 104 = 5 × 105 CFU/fly.
The instructor introduces Gram staining technique, and students
select colonies and stain them for observation under the microscope.
Allow one hour for this activity. Class results are summarized on the
board and followed with a 20–30 minute discussion.
Procedure for Instructor
In the lecture portion of the class, prior to doing the exercise in the
laboratory, the instructor should introduce the topic and assign
pertinent reading(s). This exercise has been done successfully in
three laboratory periods of two-and-a-half hours each. A suggested
timeline for the instructor is provided in Table 2.
We have done this three-week exercise with freshman students as
part of the introductory biology course at the end of the fall semester once they have been introduced to prokaryotes and cell structure. Students have been very enthusiastic about many aspects of
this exercise: using fruit flies, following a format where they make
the decision on what substance to use, a more experimental
approach, the fact that it goes over three lab periods (continuity),
as well as the relation to human health. Table 3 and Figures 2, 3,
and 4 show examples of students’ results.
Figure 1. Serial dilution example.
Table 2. Suggested schedule for instructor and/or
teaching assistant, beginning one week ahead of
Week 1 • Gather materials for fruit fly cultures
• Obtain substances unavailable to students,
Week 2 • Prepare, autoclave, and pour MRS medium
(specific for Lactobacillus)
• Autoclave pipette tips, Eppendorf microtubes,
and vials with distilled water
Week 3 • Incubate inoculated plates at 37°C for 48
hours. Keep refrigerated until needed but not
more than one week
Week 4 • Set up Gram stain stations
Table 3. Examples of students results.
Probiotic 4 × 106
Group Control 1. 8 × 106 NA
Yogurt 5. 1 × 106 One serving
Black pepper 1. 63 × 107
Corn starch 9 × 106
Group Control 9. 6 × 107 NA
Tetracycline* 1. 16 × 104 0.9 mg/mL
Penicillin 0 0.9 mg/mL
1. 65 × 108
Red food dye 4. 48 × 108 One serving
Results shown in Figures 2, 3, and 4.
Figure 2. Results from the addition of probiotic. The student
counted 40 colonies in the 10− 5 dish.