Fig 1.
Schematic of B. burgdorferi structural organization.
B. burgdorferi has a planar, flat-wave morphology that is the result of the periplasmic flagellum (PF) wrapping around the cell body between the protoplasmic cell cylinder and the outer membrane. The cross section demonstrates that the periplasmic flagella are ordered in a ribbon-like manner, wrapping around the cell body. The PFs overlap in the center of the cell. Schematic not drawn to scale.
Fig 2.
Wild-type spirochetes are more resistant to compression under external stress applied by the AFM imaging process.
Sequential tapping mode AFM topography images of (A) wild-type and (B) flaB mutant spirochetes of B. burgdorferi were taken with set point ratios of 0.8, 0.7, and 0.6. The lines indicated by various symbols in the images correspond to the height profiles (indicated by the same symbol) provided directly to the bottom of each image. Note that that the axis limits vary between the different height profiles.
Fig 3.
AFM imaging damages mutant spirochetes.
Sequential AFM images of (A) wild-type (B) flaB- mutant and (C) flgE- mutant spirochetes of B. burgdorferi demonstrating damage associated with increasing imaging forces associated with AFM. Boxes in the images indicate regions (before and after) in which permanent damage to the bacteria was observed.
Fig 4.
AFM imaging force analysis of wild-type and mutant spirochetes.
Reconstructed maximum tapping force images of B. burgdorferi flaB mutant taken at set point ratios of (A) 0.8, (B) 0.7, and (C) 0.6. The maximum force images correspond to the AFM topography images presented in Fig 2B. Below each image are representative time-resolved tip/sample forces associated with the poly-L-lysine coated mica or spirochete with arrows indicating the maximum force of each tapping event. Two oscillation cycles are shown in each force trajectory. An identical analysis was used for the wild-type (not shown). (D) The average maximum tapping force associated with imaging the mica substrate from experiments with wild-type (n = 10), flaB mutant (n = 5), and flgE mutant (n = 4) spirochetes as a function of set point ratio are shown. (E) The average maximum tapping force associated with imaging wild-type (n = 10), flaB mutant (n = 5), and flgE (n = 4) mutant spirochetes as a function of set point ratio are shown. Error bars represent the standard deviation, and # indicates p < 0.05 based on a T-test. The p values for comparing flaB- to WT were 0.01, 0.049, and 0.048 for set point ratios of 0.8, 0.7, and 0.6 respectively. The p values for comparing flgE- to WT were 0.021, 0.036, and 0.042 for set point ratios of 0.8, 0.7, and 0.6 respectively.
Fig 5.
Mechanical properties appear homogenous across spirochete bodies.
Corresponding height and reconstructed maximum tapping force images of wild-type B. burgdorferi taken at a set point ratio of 0.6.
Fig 6.
The average height of the spirochetes as a function of applied force.
RMS height of wild-type (n = 10), flaB mutant (n = 5), and flgE (n = 4) mutant spirochetes as a function of the applied maximum tapping force. Average values of RMS height are indicated by the black lines, and the standard deviation is represented by dotted lines.* indicates p < 0.01, and # indicates p < 0.05 based on a T-test.
Fig 7.
The average height of the mutant ΔW2 spirochetes as a function of applied force grown in the presence and absence of kanamycin.
RMS height of mutant ΔW2 spirochetes grown in the presence (n = 10) and absence (n = 9) as a function of the applied maximum tapping force. Average values of RMS height are indicated by the black lines, and the standard deviation is represented by dotted lines.