NADPH-cytochrome P450 reductase knockdown decreases the response to precocene I in the migratory locust Locusta migratoria.

Precocene I is a juvenile hormone antagonist that needs to be activated via oxidative biotransformation catalyzed by cytochrome P450 (CYP). NADPH-cytochrome P450 reductase (CPR) supplies CYP with electrons in the oxidation-reduction process; however, its functional role in the activation of precocene I remains unexplored. Here, the representative characteristics of CPRs were analyzed in the CPR gene of Locusta migratoria (LmCPR), the result of model docking indicated that the hydrogen bonds were formed between reduced nicotinamide adenine dinucleotide phosphate (NADPH), flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN) and NADPH-, FAD-, FMN-domains of LmCPR, respectively. Treating the fourth-instar nymphs with precocene I decreased the juvenile hormone titers of nymphs to 0.55-fold of that in acetone-treated controls, and extended the interval time between fourth- and fifth-instar nymphs. 68.75% of the treated fourth-instar nymphs developed into precocious adults in the fifth-instar. LmCPR knockdown decreased the response to precocene I in the nymphs, the occurrence rate of precocious adults induced by precocene I treatment reduced by 23.11%. Therefore, LmCPR may be involved in the activation of precocene I in L. migratoria. In addition, we generated an active recombinant LmCPR protein using a prokaryotic expression system, its activity in reducing cytochrome c was 33.13 ± 11.50 nmol CytCred/min/µg protein. This study lays the foundation for further research on the role of LmCPR in precocene I activation.

In vitro effects of caffeic acid, nortriptyline, precocene I and quercetin against Rhipicephalus annulatus (Acari: Ixodidae).

In the present study, the acaricidal effects of caffeic acid, nortriptyline, precocene I and quercetin against Rhipicephalus annulatus (syn. Boophilus annulatus) Say (Acari: Ixodidae) were evaluated. Adult immersion technique (24 ticks immersed for 2 min in one dilution of the compound) was used for the assessment of the effects of caffeic acid (0.39-100 mg/mL), nortriptyline (0.625-50 mg/L), precocene I (0.004488-5 mg/mL) and quercetin (6.25-100 mg/mL) against R. annulatus. Adult tick mortality, reproductive index, inhibition of fecundity and hatching were calculated. Caffeic acid, nortriptyline, precocene I and quercetin revealed very low adult mortality and inhibition of fecundity, even at the highest concentration tested. Quercetin (>50 mg/mL) caused blocking of hatching of eggs.

Seasonal and circadian evaluation of Ageratum conyzoides essential oil and its nematicidal activity against Caenorhabditis elegans.

The aim of this study was to evaluate the circadian and seasonal variation of Ageratum conyzoides essential oil (EO) and its nematicidal effect on the free-living nematode Caenorhabditis elegans as a model for parasitic helminths. For the seasonal study, the plants were collected from January to December 2022, at 6 a.m., and to assess the circadian rhythm, the plants were collected in April (rainy season) and October (dry season), at 6, 9, 12 a.m. and 3 and 6 p.m. The fresh plants were then subjected to hydrodistillation, and their chemical composition was analyzed using gas chromatography coupled with mass spectrometry (GC-MS). The motility test with C. elegans was carried out. The primary constituent of the oils was precocene I (65.97 to 78.42 %, respectively), followed by E-caryophyllene (6.04 to 12.16 %), comprising an average of 79.87 % of the composition throughout the year. The average yields of EOs were slightly higher in the rainy season, at 0.68 %, compared to the dry season, at 0.62 %. High light hours in the rainy season (12 a.m., 0.96 %) and in the dry season (9 a.m., 0.88 %) seem to contribute to higher daily oil yields. It was observed that the variation between the main constituents of A. conyzoides occurs in inverse proportion when analyzing the main classes of compounds present in the oils: chromenes (CH) and sesquiterpene hydrocarbons (SH). And that the month of March had the highest content of E-caryophyllene (12.16 %) when compared to the other months of the year. On the other hand, January and December had the lowest levels of precocene I (65.97 and 66.85 %). The IC50 of the EO of A. conyzoides varied according to the month and time of collection. The EO obtained in January was the most effective against C. elegans, with an IC50 of 0.01 mg/mL. Thus, A. conyzoides EO could be an alternative for nematode control, exhibiting greater efficacy if extracted during specific seasonal periods.

Anti-Ehrlichia properties of the essential oil of Ageratum conyzoides L. and its interaction with doxycycline.

Canine Monocytic Ehrlichiosis (CME) is an infectious disease caused by the rickettsia organism Ehrlichia canis which is transmitted mainly the ixodid brown dog tick Rhipicephalus sanguineus. The prevalence of E. canis infection has been increasing in recent years. The World Health Organization has been warned about antibiotics resistance and one of the way to prevent this situation is found new compound with this property. Doxycycline is the treatment of choice for this tick-borne disease. Adverse effects are noted in dogs that are sensitive to this drug. Antibiotic resistance may also occur. The present study aimed to evaluate the anti-Ehrlichia properties of the essential oil of the aerial parts of Ageratum conyzoides L. in infected DH82 cells, as well as its anti-Ehrlichia activity associated with doxycycline using the checkerboard assay. A. conyzoides is a native plant from northeast Brazil with many reports of ethnopharmacological applications. The essential oil of A. conyzoides was extracted from the aerial parts of the plant using the hydrodistillation method. E. canis-infected DH82 cells were cultured in DMEM (Dulbecco's Modified Eagle Medium), maintained at 37 °C and 5% CO2, and standardized at a 70% infection rate for the initiation of treatment protocols. The tests were first carried out with the aim of defining the IC50. The combined effect of doxycycline and A. conyzoides essential oil was then determined using the checkerboard dilution technique (checkerboard method) in which the IC50 was 200 µg/mL. The doxycycline reduction index from the combined effect was 4.90 times resulting in a synergistic effect. To the authors' knowledge, this is the first alternative treatment (alternative therapy) based on bioactive molecules that have antibacterial activity against E. canis.