## 4 Discussion
In this study, the results of the CLSM pictures indicated that thermal shock killed the bacteria in the biofilms, and this bactericidal effect was enhanced as the temperature and the exposure time increased (Figures 7 and 8). Similar trends have been observed in previous studies [39; 48]. A potential explanation for this result has been proposed in previous studies [49; 50; 51], i.e., thermal shock disrupts the hydrogen bonding and destroys the structural integrity of viral proteins such as the respiratory enzymes in the bacteria cells, and causes disintegration of the bacterial cell membrane. The results in this study agree with this explanation. In this study, two fluorescent dyes, PI and SYTO(r) 9, were used for the biofilm staining and CLSM picturing. These dyes are based on membrane integrity, i.e., SYTO(r) 9 enters all cells, fluorescing green in the CLSM, and PI enters only cells with disrupted membranes, quenching SYTO(r) 9 and fluorescing red [39]. It could be seen in the CLSM pictures that the amount of red fluorescence increased significantly as the biofilms were thermal shocked, indicating that the thermal shock killed bacteria cells by damaging the bacterial cell membranes (Figure 7). It has been reported that the strength of the biofilms would decrease when the cell viability in the biofilms reduced [52; 53; 54]. Thus, the potential mechanism for the biofilm inhibition effect in this study is that thermal shock caused disintegration of the bacterial cell membrane and killed the cells in the biofilms. As the cell viability decreased, the biofilms would become more vulnerable and detach from the solid surface, so the biomass and thermal resistance of the biofilms would decrease (Figures 3, 4 and 9). Many biofilm inhibition strategies could hardly be applied to the plate heat exchangers due to the compactness, closed internal environment of the heat exchangers, and the large flow rate and the environmental protection concerns in the TSSHPS. Thermal shocks, however, could be applied directly in the TSSHPS by stopping the treated sewage supply and pumping in hot water for a short period of time at regular intervals. This study shows that thermal shock with a certain temperature and exposure time could decrease the thermal resistance of the biofilms significantly, indicating that thermal shock is a feasible and efficient biofilm inhibition approach for the TSSHPS. To determine an appropriate thermal shock protocol for the practical TSSHPS, the influence of the temperature and the exposure time must be understood. In this study, three thermal shock protocols were tested and the amount of heat consumed in these protocols were similar. The results showed that the 80 \({}^{\circ}\)C-5 min/24 h thermal shock had the most significant biofilm inhibition effect (Figure 9), indicating that temperature has a more significant influence on the inhibiting effect than the exposure time. This could also be seen from the data fitting results of Equation (5), where \(t\) influences only the exponential term, but \(T\) influences both the exponential term and the first degree term. These resultsindicate that in practical TSSHPS, thermal shocks at higher temperatures and lower exposure times might be preferred.