## 3 Results and Discussions
### Anatomical Analysis of Leaves by Electronic Microscopy (SEM)  
Leaf stomata were observed in the different treated samples (Figure 2a,b).At the first period of treatments (t1) the stomatal density of Untreat and Treat 1 exhibited no statistical difference with a respective 671.64 and 689.55 stomata per mm\({}^{2}\) (Figure 2a), while Treat 3 showed the greatest value (794.03 stomata per mm\({}^{2}\); Figure 2b) (Table 5). At the second period (t2), again Untreat and Treat 1 showed no statistically significant difference (674.6 and 710.45 stomata per mm\({}^{2}\)), Treat 2 displayed an increase of stomata (728.36 stomata per mm\({}^{2}\)), while Treat 3 and Treat 4 recorded lower values (662.69 and 567.16 stomata per mm\({}^{2}\), respectively) in comparison with Untreat (Table 5). Comparing the treatments at t1 and t2, it can be observed that while Untreat remains substantially unchanged, Treat 1 shows a slight increase in stomatal density, while all the other treatments (Treat 2, Treat 3 and Treat 4) show a decrease in the number of stomata per unit area (Table 5). Regarding stomata diameters, higher values were measured in Untreat (19.4 \(\upmu\)m), Treat 4 (19.9 \(\upmu\)m) and Treat 3 (19.7 \(\upmu\)m) at t1, and in Treat 2 (18.5 \(\upmu\)m) and Treat 4 (18.7 \(\upmu\)m) at t2 (Table 5). At t1, Treat 3 showed higher PD/ED (1.97), while at t2 all treatments recorded a lower ratio compared to Untreat (Table 5). Two types of glandular trichomes were identified on hop leaves [38]: large peltate-type known as lupulin glands, found on the abaxial side (Figure 2c), and stalked capitate glands, called also bulbous trichomes, which are much smaller, fewer in number, and consist of a stalk and four head cells, occurring on both adaxial and abaxial surfaces, located mostly along the vessels, but rarely present on the lamina (Figure 2d black arrow). Since the latter trichomes are present in small numbers, only lupulin glands were considered in this study. At t1 all treatments showed no statistical differences with Untreat (Table 5). At t2, while Untreat remained unchanged, all treatments underwent a decrease in the density of glandular trichomes (Table 5). Two types of non-glandular hair are present on hop leaf: the predominantly cystolith-containing hairs are generally restricted to the adaxial leaf surfaces and contain calcium carbonate crystals (Figure 2e,f), as already described by [PERSON] and [PERSON] [39]; the second is the longer and rarer type of slender trichomes present only on the midribs and major veins of the abaxial leaf surfaces (Figure 2d black triangle). Since the cystoliths are more abundant and uniformly distributed, only this type was considered in this investigation. At t1, Treat 3 had the same density of Untreat, in practice (12.85 and 12.7 hairs per mm\({}^{2}\), respectively), while Treat 1, Treat 2 and Treat 4 outlined a small decrease (Table 4). At t2, Treat 2 and Treat 3 showed no statistical differences with Untreat, while Treat 1 and Treat 4 recorded the respective highest (16.84 hairs per mm\({}^{2}\)) and lowest density (7.68 hairs per mm\({}^{2}\)). The polar and equatorial diameter ratio (PD/ED) is associated with guard cell shape and is an important stomata feature. A higher ratio is correlated with an elliptic shape, characteristic of more functional stomata. Conversely, a rounded shape (smaller PD/ED) is associated with lower stomata functionality [40]. Thus, in this study, in the first period, Treat 3 recorded the best results in terms of increased number of stomata and their functionality (PD/ED ratio), while in the second period all treatments showed a worse stomata functionality, compared to Untreat.