## 3 Results and Discussion
### Effect of Microfluidic Collision on Cyst Germination Rate  
Figure 5a shows the change in the percentage of cysts relative to the total _H. lacustri_ cells during 72 h of photosynthetic cultivation in well plates. The cysts were treated under optimal microfluidic conditions (10 loops at 2.0 mL/min; see Figure 4c). The treated cultures exhibited a significantly faster cyst germination rate between 24 and 60 h compared to the untreated controls. Specifically, after 48 h of incubation, the percentage of cysts in the treated cultures decreased to 18.5 \(\pm\) 6.1%, whereas the untreated controls maintained a higher cyst percentage of 33.0 \(\pm\) 5.2%. This accelerated germination process was further supported by the high-speed footage captured from 12 to 72 h for both control and treated cultures (see Supplementary Video S1). We hypothesize that the increased cyst germination  
Figure 4: Effect of microfluidic loop number on **(a)** cell viability and **(b)** reactive oxygen species levels in _Haematococcus lacustri_ cysts. **(c)** Time course profile of total cell number density for 72 h of photosynthetic well plate cultivation using cysts treated with varying loop numbers (3, 6, 10, 20, and 30). Error bars represent the mean and standard deviation of three independent experiments (\(n\) = 6). Different italicized letters (\(a\), \(b\), \(ab\), and \(c\)) indicate the significance of the differences (\(p\) < 0.05). DCF: 2′.7′-dichlorofluorescein. n.s. denotes non-significance between the groups. efficiency is associated with the enhanced total cell density observed after 72 h, as shown in Figure 4c. This suggests that microfluid collision treatment stimulates the transition from dormant cysts to actively dividing cells and also supports sustained cell growth. During the cyst germination of _H. lacustris_, as depicted in Figure 2, a single cyst cell undergoes repeated mitotic events and cytokinesis, resulting in 2\({}^{\text{n}}\) daughter cells (e.g., 2, 4, 8, and 16). To examine the impact of microfluidic treatment on this germination pattern, microscopic images were analyzed at the 48 h mark, considering the significant differences observed between treated and control cultures (Figure 5b). Notably, early germinating stages producing two and four daughter cells were absent, indicating advanced progression in germination events, regardless of microfluidic treatment. Two distinct responses were observed. In untreated cells and those exposed to lower loop numbers (three and six), the proportion of germinating cells producing eight daughter cells was substantially lower, at 56.8-62.5%, compared to the cultures subjected to higher loop numbers (10 to 30), which exhibited ratios of 81.3-81.8%. The data from Figure 5a,b collectively suggest that mechanical collisions influence both the morphology and physiological cell division patterns of _H. lacustris_ cysts. Figure 5: **(a)** Time course profile of the cyst cell ratio relative to total cell count during 72 h of photosynthetic well plate cultivation of _Haematococcus lacustris_ using cysts treated with the optimal microfluidic treatment conditions (10 loops at 2.0 mL/min). **(b)** Ratio of germinating cell types producing 8 and 16 daughter cells after 48 h of well plate incubation with cysts treated at different microfluidic loop numbers (3, 6, 10, 20, and 30). Refer to Figure 2 for representative microscopic images of germinating cells. Each sample includes at least 100 randomly selected cells from three independent experiments (\(n=6\)). Different italicized letters (\(a\), \(ab\), \(b\), \(c\), \(d\), \(e\), and \(f\)) indicate the significance of the differences (\(p<0.01\)). n.s. denotes non-significance between the groups. The increase in cell density may be linked to the activation of mechanoreceptors in the cell wall that induce cell growth [52]. The cell membrane plays a crucial role in various cellular activities, such as activating ion channels and maintaining intracellular ion balance that are vital for cell resistance to mechanical damage.