Real-Time Monitoring of <i>Streptococcus mutans</i> Biofilm Formation Using a Quartz Crystal Microbalance

Abstract

The ability of Streptococcus mutans, a well-known etiological agent in dental caries, to attach and form a biofilm is an important key to its virulence. The effects of various environmental factors (i.e. sucrose concentration, flow rate and temperature as well as genetic manipulations) on the capability of S. mutans (UA 140) to attach, form and detach were monitored in situ using quartz crystal microbalance. The biofilm growth rate was much slower than that of planktonic growth. Greater availability of sucrose contributed to biofilms with less lag time, lower doubling times and earlier detachment. Flow rate experiments showed that as the shear stress was reduced, the maximum mass accumulated also decreased. However, the detachment process was independent of shear force, perhaps indicative of quorum sensing. Increasing the incubation temperature from 37 to 40°C extended the lag period and inhibited the ability of the biofilm to attach readily. Absence of either the ciaH, luxS, gtfB or gtfC genes also greatly affected the ability of the S. mutans to adhere to a surface in comparison to the wild type. Quartz crystal microbalance results indicate that the gtfC gene possibly has a greater contribution to biofilm attachment than the gtfB gene, that the presence of the luxS gene is critical for attachment and that the ciaH gene primarily affects the initial reversible attachment of the biofilm.

Towards real-time observation of conditioning film and early biofilm formation under laminar flow conditions using a quartz crystal microbalance

“…8). This agrees with the findings of a previous study [45] where there was a long lag time of early biofilm formation under nutrient-limited growth conditions. The long lag time of biofilm formation in a mineral salts/trace elements-limited growth medium may, in part, be caused by the effect of the bulk ionic strength of the aqueous environment: at low bulk ionic strengths (generally I < 0.001 M), bacterial attachment is inhibited by DLVO-type electrostatic repulsion, but at high bulk ionic strengths (generally I > 0.1 M), there will be no net potential energy barrier to contact between the bacteria and surface [23,46].…”

luxS-Based Quorum-Sensing Signaling Affects Biofilm Formation in <i>Streptococcus mutans</i>

“…Recently, a different methodology, using a quartz crystal microbalance (QCM), has been reported to observe the real-time biofilm growth characteristics of the luxS mutant of S. mutans [Tam et al, 2007]. The QCM approach correlates the quartz crystal resonance frequency changes to the biofilm actual mass deposited on the surface.…”

“…The QCM approach correlates the quartz crystal resonance frequency changes to the biofilm actual mass deposited on the surface. Tam et al [2007] reported that the deletion of the luxS gene inhibited the ability of the bacteria to attach, although the mutant grew almost as well as the wild-type. The different results of the present research and QCM might be due to different research focuses of the two experiments of QCM and BioFilm Ring Test (QCM measured the mass of biofilm, however the BioFilm Ring Test measured the immobilization force of the beads due to the viscoelastic resistance of the biofilm matrix, which can immobilize paramagnetic beads in the magnetic field).…”

Pronounced Effect of the Nature of the Inoculum on Biofilm Development in Flow Systems

“…Biofilm growth rates have been determined by various techniques, such as fluorescence in situ hybridization (FISH) (12,31), measuring the incorporation of radioactive substances like [ 3 H]thymidine and 32 P (8, 9), microscopy (3,13,17), measuring the total increase in biofilm mass (both cells and extracellular polymeric substances) (24,28), colorimetric 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenyl-amino)carbonyl]-2H-tetrazolium hydroxide (XTT) assays (26), or measurement of amide II bands, as determined by attenuated total reflectanceFourier transform infrared spectroscopy (6). Some of the above-mentioned techniques suffer the drawback that the bio-films have to be sacrificed with sampling or that the measured increases do not distinguish between live and dead matter in the biofilm (i.e., increases measured might not represent an accurate increase in viable cell numbers).…”