The role of the pH conditions of growth on the bioadhesion of individual and lawns of pathogenic Listeria monocytogenes cells

Abstract

The work of adhesion that governs the interactions between pathogenic Listeria monocytogenes and silicon nitride in water was probed for individual cells using atomic force microscopy and for lawns of cells using contact angle measurements combined with a thermodynamic-based harmonic mean model. The work of adhesion was probed for cells cultured under variable pH conditions of growth that ranged from pH 5 to pH 9. Our results indicated that L. monocytogenes cells survived and adapted well to the chemical stresses applied. For all pH conditions investigated, a transition was observed in the generation time, physiochemical properties, biopolymer grafting density and bioadhesion for cells cultured in media adjusted to pH 7 of growth. In media with pH 7, the generation time for the bacterial cells was lowest, the specific growth rate constant was highest, the cells were the most polar, cells displayed the highest grafting density of surface biopolymers and the highest bioadhesion to silicon nitride in water represented in terms of the work of adhesion. When compared, the work of adhesion values quantified between silicon nitride and lawns of L. monocytogenes cells were linearly correlated with the work of adhesion values quantified between silicon nitride and individual L. monocytogenes cells.

Figure 1

Tapping mode images of L. monocytogenes cells grown in BHIB adjusted to different pH values and cultivated in their late exponential phase of growth. Images captured in pH 6 and in pH 9 are 10 × 10 μm while other images are 20 × 20 μm. Heights in all images are 600 nm. The means of heights of 17 cells grown under each investigated pH condition were 341 ± 40, 378 ± 52, 377 ± 55, 376 ± 49 and 307 ± 46 nm in pH 5, 6, 7, 8 and 9 respectively. The error represents the standard deviation.

Figure 2

Example of an AFM retraction curve measured between a silicon nitride AFM cantilever and L. monocytogenes surface biopolymers in water. The gray shadowed area represents the work of adhesion or adhesion energy in fJ. The black arrows at the top of the curve indicate the bounds of integration where the adhesion energy was computed using equations 1 and 2.

Figure 3

Force-distance approach curves measured between L. monocytogenes grown under variable pH conditions and silicon nitride in water. Each curve is the average of 225 individual curves measured on 15 individual cells taken from three different cultures. The solid black lines are the steric model (eq. 4) fits to the data. Values of L and Γ used in fitting the curves above are given in Table 2. The ability of the steric model (eq. 4) to fit the data was judged based on the estimated values of r² (the coefficient of determination, often used to judge the adequacy of a regression model) using the TableCurve fitting program (Windows, version 1.11, Jandel Scientific).

Figure 4

Histograms showing the distribution of the work of adhesion affinities measured between individual L. monocytogenes surface biopolymers grown at (A) pH 5, (B) pH 6, (C) pH 7, (D) pH 8, and (E) pH 9 and silicon nitride in water. The probabilities of occurrences of the work of adhesion values were normalized by the total number of work of adhesion events measured for each investigated pH condition. Solid lines indicate the log-normal dynamic peak function with four parameters fits to the data.

Figure 5

A scatter graph that shows the relationship between the pH of growth of L. monocytogenes and the mean of the work of adhesion quantified between individual or lawns of L. monocytogenes cells and silicon nitride in water using AFM and the macroscale HM thermodynamic-based model, respectively.

Figure 6

A comparison of the work of adhesion measured by AFM and the macroscale work of adhesion calculated from the contact angle measurements using the thermodynamic-based HM model. The solid line represents the linear fit to the data with the formula $(W_{\mathit{adh}}HM(fJ)=1.121\times W_{\mathit{adh}}\mathit{AFM}(fJ)+0.288,r^2=0.923)$.