Battle of the Bacteria: Characterizing the Evolutionary Advantage of Stationary Phase Growth †

Timeline for the four experiments in the GASP module. The module was implemented in a lab course that met twice a week (Lab Period A and B). Lab Period A was three hours in duration while Period B was one hour. Each activity during lab is associated with one of the four specific experiments. (1) Growth of cells into long-term stationary phase and GASP assay, (2) Examination of RpoS activity, (3) Cell stress measurement, (4) Examination of mutation frequency. Specific times required for each activity are indicated on the figure in minutes.

Examples of student data illustrating the GASP phenotype. (A) Dilution plating on selective media. Cells are from the co-culture experiment (started in week three of the module). Day 0 refers to the initial culture inoculation and day 7 refers to one week of co-culture growth. Aged cells are distinguished from un-aged cells by the presence of distinct antibiotic resistance markers present in each strain. Cultures are plated with ten-fold serial dilutions. (B) Colony forming units (CFU) per ml of culture are calculated from the dilution plates and graphed on a plot of CFU/ml versus time. The GASP phenotype is illustrated by greater survival values of aged cells over time compared to the un-aged population in the culture. Growth in different types of media can impact the GASP phenotype.

Assessments confirm that students achieved the module learning objectives. (A) A 12-question pre-/post-test was administered before and after the GASP module in laboratory sections during Fall quarter 2014 and Winter quarter 2015 (n = 197 students combined). Performance on each question (Q) and the corresponding learning objective (LO) it assessed is indicated. All post-test gains are statistically significant (p < 0.001 by t-test) with the exception of question 4. (B) Students noted their agreement with the indicated statements on a 5 -point Likert scale (5 = strongly agree, 1 = strongly disagree) upon completion of the module. They were asked to state their current agreement and provide a retroactive agreement (the pre-module data). Post-test gains are statistically significant (p < 0.001 by chi-square test). Questions were asked using the iClicker response system (n = 190 students). (C) Student performance on the pipetting test before and after module completion (n = 198 students). Passing refers to whether a student’s dilution fell within 0.050 OD595 units from a standard value obtained by the course instructor who performed the same dilutions multiple times. The number of students who passed post-module was significantly greater than pre-module (p < 0.001 by t-test). (D) Student performance on exam questions categorized by whether or not they were GASP module-specific and by Bloom’s level (Bloom’s 2 versus 3, 4, and 5) (n = 201 students). The difference between GASP and Other questions of Bloom’s level 3, 4, and 5 was statistically significant (p < 0.001 by t-test). Question numbers in each category were GASP (BL2) = 5, Other (BL2) = 10, GASP (BL3, 4, 5) = 19, Other (BL3, 4, 5) = 38.