1. Stochastic Receptor Expression Allows Sensitive Bacteria to Evade Phage Attack. Part I: Experiments 
E. Chapman-McQuiston, X.L. Wu 
Biophysical Journal 
Volume 94, Issue 11, Pages (June 2008)
DOI: /biophysj
Copyright © 2008 The Biophysical Society Terms and Conditions

2. Figure 1
Epifluorescent microscopy of λ-phage bound to bacteria Ymel. The λ-phage are labeled with AlexaFluor-488 and incubated with the bacteria for 40min (see details in the Methods). The two pictures (a and b) are identical except that the intensity gain is different. Typical bacteria adsorb 200–300 phage particles, yielding brightened cell envelopes. However, a minority of bacteria adsorb a small number of phage particles and can be seen only when a significant gain is applied to the CCD camera as delineated by the encircled cell in b. For bacteria with a small number of LamB receptors, the attached phage can be counted because they appear as individual diffraction-limited spots on the bacterium.
Biophysical Journal  , DOI: ( /biophysj )
Copyright © 2008 The Biophysical Society Terms and Conditions

3. Figure 2
(a) Total fluorescence readings at different dilutions were taken from samples containing phage (squares), fluorescent latex spheres (circles), Ymel + phage complexes (diamonds), induced LE392 + phage complexes (triangles), and the cell background (stars). After a background subtraction, the dilution curve was fit with a straight line with a zero intercept. These values were used to calculate the average receptor number per bacterium as shown in Table 1. (b) The phage depletion rate θ (≡ ln((P(t)/P(0))/B(0)) is measured for Ymel (squares) and for induced LE392 (triangle). The measurements were carried at 37°C in the λ-dilution buffer supplemented with 10mM of MgSO4. The initial bacterial and phage concentrations were S(0)≈108cm−3 and P(0)≈5×104cm−3. As can be seen, the adsorption is more rapid with LE392 cells than with Ymel cells because of the difference in the mean receptor number. (c) The FI of different bacterial preparations was characterized by flow cytometry. The displayed curves include Ymel cells without labeling (black), uninduced LE392 with labeling (blue), Ymel with labeling (red), and induced LE392 with labeling (purple). The vertical arrows indicate secondary bumps that are often seen in the flow cytometry data. (d) After deconvolution using the cell background data, the number of phage attached to each bacterium can be calculated, and the resulting receptor PDFs are given for the uninduced LE392 (blue), the Ymel (red), and the induced LE392 (purple) bacteria. The binned PDFs are constructed based on measurements of more than 5×104 cells.
Biophysical Journal  , DOI: ( /biophysj )
Copyright © 2008 The Biophysical Society Terms and Conditions

4. Figure 3
The response of Ymel (a), induced LE392 (b), and uninduced LE392 (c) bacterial populations to perturbations by different phage pressures P(0). The phage concentrations, P(0), for runs in a and b are similar for the corresponding colors, and the detail numbers are given in the legend of b. The phage concentrations for runs in c are also given in the legend of the figure. For both bacterial strains in a and b, three different initial responses were observed, corresponding to growing, decaying, or having a zero growth rate. After killing with a high P(0), the subsequent bacterial growth rate is often very low immediately after killing and persists for several hours. Exponential growth (not displayed) is always observed in long times. As shown in c, the uninduced LE392 cells are much less affected by increasing P(0). A close inspection of the earlier data, which are displayed in the inset, the growth rates for the two highest P(0)=2×109cm−3 (red) and 1010cm−3 (black) are nearly identical.
Biophysical Journal  , DOI: ( /biophysj )
Copyright © 2008 The Biophysical Society Terms and Conditions

5. Figure 4
(a) Simultaneous measurements of bacterial (LE392) and phage concentrations as a function of time. Here the initial MOI=3 is low so that the phage concentration decreases with time. However, the combination of killing of the bacteria and reproduction of phage at t=1h significantly increases MOI to ∼100. The net result is that the constant-P assumption is satisfied for t>1.5h as shown. (b) Killing curves in a reduced-magnesium-salt medium. To determine whether the initial adsorption is responsible for the observed killing curves, the measurements were also conducted with Ymel in the λ-buffer with reduced MgSO4 (1mM instead of 10mM). In the reduced-salt medium, the phage adsorption rate was decreased by a factor of four. The bacterial concentration was S(0)=2×106cm−3. A range of phage concentrations was used, and they are labeled in the figure. The solid lines are a guide to the eye.
Biophysical Journal  , DOI: ( /biophysj )
Copyright © 2008 The Biophysical Society Terms and Conditions

6. Figure 5
(a) Killing curves with high phage concentrations. The measurements were carried out with P(0)=4×1010cm−3. Two different bacteria, Ymel (circles) and Ymel + pTAS1 (squares), were used. To have significant colony counts, the bacterial concentration B(0) was increased (3×107 and 2×108cm−3) in this set of measurements. The cells with the plasmid showed a slightly faster killing and a lower level of survival bacteria at long times. The inset delineates in more detail the earlier killing behavior. After ∼5h both cells begin to grow exponentially, as indicated by the straight lines. The low intercept values at t=0 indicate possible contributions by mutants. (b) The cells were tested for their sensitivity to phage infection. After ∼6h, the fraction of sensitive cells, S(t), diminishes in the total cell population, B(t). Here squares are for Ymel + pTAS1, and the circle is for Ymel.
Biophysical Journal  , DOI: ( /biophysj )
Copyright © 2008 The Biophysical Society Terms and Conditions