Tricomoníase aviária: eficácia de óleos essenciais e nanoemulsões in vitro e in vivo

Abstract

Avian trichomoniasis is a parasitic disease caused by the protozoan Trichomonas gallinae. This parasite most commonly affects chicks and may be responsible for disease outbreaks in bird populations. The treatment for this disease is carried out with metronidazole (MTZ). The use of essential oils (EOs) from plants represents a source of potential active principles. In this context, this study aimed to evaluate the effectiveness of essential oils and nanoemulsions through in vitro and in vivo activities on T. gallinae trophozoites. To evaluate the susceptibility of essential oils of Eucalyptus (Eucalyptus globulus) and Basil (Ocimum basilicum) and their respectives nanoemulsions, sterile 96-well-plates were used to incubate the trophozoites with different concentrations. The parasites were seed at an initial density of 1x10 6 trophozoites/mL and incubated with the oils and nanoemulsions. Four controls were performed: a) trophozoites only (negative control), b) trophozoites plus 100µM metronidazole (as a positive control) c) trophozoites plus the vehicle for solubilization of derivatives (0.01% Tween) and d) white nanoemulsion. To verify the viability of trophozoites, it was observed the motility, morphology and exclusion by Tripan Blue dye (0.4%). The data obtained by analysis of screening compounds showed that 1.5% of O. basilicum EO and 1.25% of nanoemulsion reduces 100% of trophozoite viability when compared to the negative control. EO E. globulus nanoemulsion 1.5% and 2% essential oil. Controls CN (negative control), TW (tween 0.01%) and NB (blank nanoemulsion) exhibited positive motility and negative staining with trypan blue, while the MTZ control reduced parasite viability by 100% after 24 hours of exposure. The data obtained in this study indicate that EOs and nanoemulsions can contribute as effective agents in the control of T. gallinae infections. In vitro and in vivo assay were realized with essential oil (EO-CL) and nanoemulsion (N-CL) of lemongrass (Cymbopogon flexuosus). In vitro assays were performed with 1x10 6 trophozoites/mL and EO-CL and N-CL at concentrations: 110, 220, 330, 440, 550, 660, 770 and 880 and four controls: CN (culture medium and trophozoites), MTZ (trophozoites plus 800 of metronidazole), TW (trophozoites plus vehicles used for solubilization of derivatives (0.01% Tween) and NB 9

(nanoemulsion blank 880 ). In vivo assay was carried out on 35 quails (Coturnix coturnix) experimentally infected with 4x10

4 trophozoites mg/kg and were divided into seven groups (n = 5): A (control - healthy), B (infected control), C (control TW 0 .01%), D (NB 0.88 mg/kg), E (MTZ drug 25 mg/kg, F (OE-CL at 0.55 mg/kg) and G (N-CL at 0.44 mg/kg) and were treated for 7 consecutive days. In vitro test showed that concentrations of OE-CL (550 g/ml) and N-CL (440 g/ml) reduces 100% the of trophozoites viability. At in vivo test, OE-CL was effective on the 4th day of treatment and N-CL after the 3rd day, and MTZ at the therapeutic concentration was effective on the 7th day. To evaluate the in vitro cytotoxicity, antioxidant and antiparasitic activity of citral (3,7-Dimethyl-2,6-octadienal) and geraniol (trans-3,7-Dimethyl-2,6-octadien-1ol), majority compounds present in lemongrass, the

in vitro assays were performed with 1x106 trophozoites/mL and citral and geraniol at concentrations of 10, 20, 30, 40, 50, 60, 70, 80, 90 and 100 M, and four controls: CN (culture medium and trophozoites), MTZ (trophozoites plus 100 M metronidazole), TW (trophozoites plus vehicles used for solubilization of derivatives (0.01% Tween). In the cytotoxicity test, citral have promoted the determination of reactive oxygen species (ROS) and the levels of thiobarbituric acid reactive species (TBARS) showed that citral and geraniol have an antioxidant activity. In vitro test showed that concentrations of citral (60 M) and geraniol (50 M) reduced 100% of trophozoites viability. In the cytotoxicity test, geraniol did not cause effects and damage to VERO cells in 24 hours of time-dependent treatment, when compared to the control, the determination of the levels of ROS and TBARS showed that citral and geraniol have an antioxidant activity. The molecular docking experiment demonstrated that there may be a potential for citral and geraniol to inhibit a hydrogen of some crucial enzyme for the survival of T. gallinae. In order to measure the in vivo effect of essential oil and nanoemulsion of Cymbopogon flexuosus in positive quails, through determination of oxidative stress parameters, histological analysis of organs such as liver, kidney and digestive system mucosa and biochemical analyzes were performed. The tests realized to determine the levels of ROS, TBARS and Superoxide dismutase (SOD), showed that EOs and nanoemulsion has antioxidant activity. No histological changes were observed in birds infected in the oral cavity, liver and kidneys, however, they presented lesions in the esophagus.. The data obtained in this study indicate that EOs and nanoemulsions can contribute as effective agents in the control of infections, also showing C. flexuosus as a potent natural and effective antitrichomonal agent against T. gallinae.