## 3 Results and Discussion
### Bacterial Inactivation
#### 3.3.2 Inactivation of Bacteria in a Flow Reactor  
The assessment of river water for bacterial counts (Section 3.3.1) provided sufficient information to plan experiments on bacterial inactivation in the flow reactor. The initial bacterial load was in a realistic range of 1200 to 54,000 CFU/L (Figure 8a). Figure 8a shows the changes in the number of bacteria at the initial stage of inactivation (7 min). Figure 8b,c show the relative survival of bacteria (N/N\({}_{0}\)) over the full time range (28 min). At a low initial bacterial load (1200-6900 CFU/L), a rapid and significant reduction in bacterial counts is observed within the first minute of contact (Figure 8a). The relative survival of bacteria also decreases rapidly within the first minute of inactivation (Figure 8b). After 10 min, the bacteria are almost completely inactivated. However, bacteria survive better at higher initial concentrations (Figure 8c). At high initial bacterial loads (\(\geq\)900 CFU/L), bacterial inactivation becomes slow and incomplete (Figure 8a). Even after 28 min, a significant portion of the bacterial cells remain viable, especially at N\({}_{0}\) > 40,000 CFU/L (Figure 8c). There is a clear concentration dependence: the higher the initial bacterial count, the less effective the inactivation (Figure 8c). Figure 8b,c clearly show that the kinetics of bacterial inactivation are highly nonlinear. Bacterial inactivation appears to occur in two stages: first, there is a rapid initial decline in bacterial numbers, followed by a slower second stage. There are several possible causes for the slow inactivation phase. The obvious cause is the increasing number of dead bacteria, whose damaged cells lose their internal cytoplasmic components. The washed-out molecules react with hydroxyl radicals, which reduces the number of active radicals and slows down the rate of bacterial inactivation. Another reason for the slowdown in bacterial inactivation is the fact that the bacterial population includes bacteria with varying resistance to aggressive radicals. Some bacteria have better defense mechanisms and can reproduce within about 30 min of the experiment, resulting in a lower inactivation rate. An additional inhibiting effect may occur as a result of the formation of a biofilm on the catalyst surface. Figure 9 illustrates how increasing the bacterial concentration facilitates biofilm formation in the catalyst bed. At low bacterial concentrations (left side of Figure 9), the catalyst surface remains clean and exposed to hydrogen peroxide, allowing continuous formation of reactive oxygen species. The obvious effect is poor bacterial survival. In contrast, at higher bacterial concentrations (right side of Figure 9), bacterial cells colonize the catalyst surface and begin forming a protective biofilm matrix. This biofilm is mainly composed of polysaccharides, proteins, and extracellular nucleic acids [33,34]. These extracellular polymeric substances significantly increase the adhesion of bacterial cells to the catalyst surface. The biofilm progressively envelops the catalyst surface, preventing its contact with hydrogen peroxide, thus reducing the number of radicals and facilitating bacterial survival. Similar protective mechanisms have been observed in _Escherichia coli_ and _Pseudomonas aeruginosa_, where shielding with extracellular polymeric substances increases the survival of bacteria under conditions of oxidative and chemical stress [33]. This survival strategy in various Gram-negative bacteria is controlled by chemosensory signaling systems, allowing them to adapt to environmental stresses [34,35]. Biofilm-based shielding may explain the observed stabilization of bacterial viability at higher initial bacterial counts. A similar phenomenon of physical covering of the catalyst surface with a dense biofilm is known as biofouling [36,37]. Therefore, the formation of bacterial biofilms on the catalyst surface may represent a synergistic effect: improving microbial persistence while simultaneously reducing ROS formation.