## 5 Discussion  
This in vitro study investigated the effects of professional oral hygiene treatments on surface microtopography and microbial biofilm adhesion on zirconia obtained by different techniques (CAD/CAM milled and veenered). Both forms of zirconia had surfaces treated differently: glazed and polished. The results of this study showed that the null hypothesis was rejected, because the microtopography of the sample surfaces and biofilm formations were different, depending on the oral hygiene maintenance treatment. In this research, ultrasonic scaling and brushing were performed to simulate professional plaque control. These methods belong to the mandatory and recommended oral hygiene maintenance procedure after prosthetic treatment. There is the recommended control and the performing of routine oral hygiene procedures to be carried out at least twice a year after prosthetic and periodontal surgical treatment [24]. Previously reported data [21] showed that the the time required for the treatment of one tooth with ultrasonic scaler during periodontal therapy ranges from 0.35 to 3.90 min. Based on this study, ultrasonic scaling was performed for 1 min and repeated 10 times, imitating a procedure carried out over 5 years. The microtopography properties of the tested zirconia samples were qualitatively evaluated by SEM. Observation with a SEM microscope was performed at 150\(\times\) magnification. This magnification allowed us to see comprehensive changes on the surfaces of the tested samples. Other research performed the analysis at a higher magnification [25; 26; 27]. In this case, higher magnification would not show the changes on the surfaces caused by the applied ultrasonic scaling and brushing techniques described in this experiment in such an obvious and striking way. In this study, the biggest surface defect was caused by the ultrasonic scaling treatment, especially on the glazed surfaces of the zirconia samples, which could be explained by the glassy structure of the glaze, which was literally broken up by ultrasound vibrations. Some studies have showed a significant increase in surface roughness after scaling [24; 28], while others have not [27]. This diversity could be caused by different timings of ultrasonic scaling in previous studies and the differently treated surfaces and types of ceramics. Disruption of the surface topography by brushing was reported in other studies [26; 29; 30], but in this study, it was particularly pronounced in the glazed zirconia samples. Damage caused by brushing was less but not insignificant, especially when we knew that the microtopography of the ceramic restorations surface was related to the formation of the biofilm [31; 32]. Biofilm has several stages in its formation. It represents dense microcommunities, which have the ability to adapt to changes in the environment by changing their gene expression patterns. The formation and composition of the biofilm is influenced by the method of adhesion to the surface. Each microorganism has its own unique method (binding using flagella, saws, proteins or polysaccharides). The initial phase of attachment of microorganisms is the key phase for the formation of a biofilm, and this process can further go in two directions: the microorganisms can continue to stick to the surface and to each other, or they can return to their free form (planktonic). It could be said that microorganisms have created a unique way to survive by forming a biofilm [33]. It has been proven that a biofilm is formed on all surfaces in the oral cavity, both natural tissues and artificial materials. It is the microorganisms from the biofilm with their products that affect the inflammatory processes in the mouth and the durability of dental restorations. Scanning electron microscopy (SEM) showed a significant difference in biofilm formation on different materials and tooth enamel, and this was related to the roughness of the surfaces of the examined materials [34]. Biofilm adhesion is influenced by several factors.