From the data, the following parameters were determined: time-to-peak heat flow, peak heat flow, mean heat flow (that is, heat flow averaged over the period, 72–420 h, as during this time it stayed on a stable plateau level after reaching the peak heat flow), and total heat produced up to 480 h (computed as the integral of the heat-flow-vs.-time curve between 0 and 480 h). For each of the four IMC parameters determined, the median, mean, and standard deviation were
Epigenetic inhibitor mw computed. Test of significance was conducted using the Kruskal–Wallis analysis of variance method and a commercially available software package (stata Statistical Software, release 16; StataCorp, College Station, TX). Significance was denoted at P < 0.05. SEM revealed that a biofilm was present over the entire surface of a protein-coated titanium disk (Fig. 1a), although minor variations were observed in the density and areas covered by the EPS matrix (Fig. 1b). FISH/CLSM showed bundles of F. nucleatum cells that formed the framework of the biofilm in which both coccal species were observed to bind (Fig. 2). Hardly, any co-adherence between the two coccal learn more species was detected. While the relative proportions of the bacterial species stayed unchaged within the biofilms, the total bacterial count showed significant differences
(P < 0.05); however, the biological meaning of the minor differences in total counts is doubtful (Table 2). A typical IMC heat-flow-vs.-time plot is given in Fig. 3. In 17 of 24 samples, after the heat flow reached a peak, it gradually reduced and finally returned to baseline value. In each of the other seven samples, however, after the heat flow reached a peak, it did not reduce to baseline value but remained at a high plateau. While the variability in the time-to-reach Mirabegron peak, heat flow is low, each of the other three parameters determined showed large variability (Table 3). Microscopic analyses have proven to
be invaluable tools in describing biofilms in terms of their structure and association with a surface; however, no real-time information about the dynamics of the metabolic activity and biomass formation can be obtained. The present study is the first of its kind characterizing a multispecies in vitro biofilm using both well-established microscopic methods (SEM and FISH/CLSM) and a very sensitive calorimetric method (IMC). Imaging by SEM was used to gain the first overview of the formed biofilm. As intended, the scans revealed biofilms that covered the entire surface with bacteria partially embedded in EPS, indicating that 0.2% glucose (Tenuta et al., 2006; Filoche et al., 2007) was sufficient to induce EPS formation. In addition, SEM showed that F. nucleatum served as the central ‘bridging organism’ or framework in the biofilm architecture, demonstrating inter- and intraspecies cellular binding (Lancy et al., 1983; Kaplan et al., 2009; Merritt et al., 2009).