In vitro and ex vivo anti-Pythium insidiosum potential of ozonated sunflower oil

This study sought to evaluate the in vitro and ex vivo susceptibility of Pythium insidiosum to ozonized sunflower oil (OSO) and verify the morphological alterations of OSO-exposed hyphae. Susceptibility assays were performed according to the broth microdilution protocol M38-A2/CLSI, and the minimal inhibitory (MIC) and minimal oomicidal (MOC) concentrations were also determined. Non-ozonated sunflower oil (SO) was used as the oil control. Additionally, kunkers from equine pythiosis were exposed to OSO. Damages caused by OSO and SO on P. insidiosum hyphae ultrastructure were verified using scanning electron microscopy. The MIC range for OSO was 7000 to 437.5 mg/mL, and the values for SO were higher, ranging from 56000 to 14000 mg/mL. The MOC was equal to MIC for both oil formulations. The OSO fully inhibited the oomycete growth from kunkers, although there was P. insidiosum growth in the kunker control in 24 h of incubation. The SEM analyses showed that both OSO and SO caused morphological alterations in P. insidiosum hyphae, highlighting the presence of cavitation along the hyphae with loss of continuity of the cell wall, which was more evident in the OSO-treated hyphae. The OSO had the best oomicidal activity, leading us to believe that our findings may support future research containing this formulation to be applied in integrative medicine protocols to control pythiosis in animals and humans.