ABSTRACT
Bovine Trichomonosis, an endemic sexually transmitted disease in countries with extensive livestock and natural service, represents one of the most common causes of reproductive failure. 5-nitroimidazoles and their derivatives are used for its treatment, mainly metronidazole (Mz). The emergence of resistance mechanisms adopted by the parasites against the drug and failure of the treatments suggest the need to investigate susceptibility and obtain current values. The available information of in vitro susceptibility of these drugs comes from the use of a diversity of methodologies and criteria, especially observation of the mobility of the parasite under the optical microscope to evaluate its viability. These techniques are arduous and time consuming and lead to a subjective assessment dependent on the operator, the methodology used, and the morphology adopted by the protozoan. In this sense, flow cytometry has proven to be a fast and efficient method to evaluate viability in other protozoa. The aim of this study was to evaluate the in vitro susceptibility of six bovine isolates of Tritrichomonas foetus to Mz in aerobic (AC) and anaerobic (ANC) conditions by means of the calculation of the 50% inhibitory concentration (IC50), by flow cytometry, and also to analyze minimum lethal concentration (MLC) by means of recovery tests post-treatment in vitro. IC50 values ranged from 1.06 to 1.25 µM in ANC and from 1.44 to 3.03 µM in AC, these being the only ones reported at 48 h for these protozoa. With respect to MLC at 48 h, the results were from 3.67 to 7.35 µM in ANC, and from 7.35 to 14.7 µM for AC, where two isolates (Tf0 and Tf2) for AC and one (Tf2) for ANC showed higher values than those described in the literature. Flow cytometry has proven to be an effective, rapid and objective methodology and very useful in susceptibility tests. The data obtained through these tests allow us to describe the susceptibility exhibited by these protozoa, this being valuable information when establishing dosages in Mz treatments.
Subject(s)
Flow Cytometry , Metronidazole/pharmacology , Parasitic Sensitivity Tests , Tritrichomonas foetus/drug effects , Animals , Cattle/parasitology , Inhibitory Concentration 50 , Nitroimidazoles/pharmacologyABSTRACT
BACKGROUND: Tritrichomonas foetus is the etiologic agent of the sexually transmitted disease Bovine Trichomonosis (BT). In Argentina, BT is endemic and represents a relevant health problem that causes reproductive inefficiency in cattle and large economic losses. Metronidazole is the drug of choice in the treatment of BT. Treatment has been associated with a temporary resolution of the clinical signs but is not able to control the disease. In recent years, the apparition of in vivo and in vitro aerobic and anaerobic resistance leading to ineffective treatments has been reported. AIMS: Thus, the aim of the present study was to explore the susceptibility of six different isolates of T. foetus under aerobic (AC) and anaerobic (ANC) conditions. RESULTS AND DISCUSSION: Six isolates of T. foetus were obtained from samples of preputial smegma of bovine origin. Values of minimum lethal concentration and minimum inhibitory concentration were higher than those observed in other works and represent current data in Argentina and provide information to establish new treatment protocols.
Subject(s)
Antiprotozoal Agents/pharmacology , Metronidazole/pharmacology , Tritrichomonas foetus/drug effects , Aerobiosis , Anaerobiosis , Animals , Argentina , Cattle , Cattle Diseases/parasitology , Protozoan Infections, Animal , Trichomonas Infections/veterinary , Tritrichomonas foetus/isolation & purificationABSTRACT
The objective of this study was to elucidate the signalling pathways initiated by cAMP once inside the Xenopus laevis oocyte, where it triggers and maintains vitellogenin endocytic uptake. Our results showed the presence of Xepac transcripts at all stages of oogenesis and we demonstrated that a cAMP analogue that exclusively activates Xepac, 8-CPT, was able to rescue the endocytic activity in oocytes with uncoupled gap junctions. Inhibition experiments for the IP3/Ca2+ signalling pathway showed either a complete inhibition or a significant reduction of the vitellogenic process. These results were confirmed with the rescue capability of the A-23187 ionophore in those oocyte batches in which the IP3/Ca2+ pathway was inhibited. Taking our findings into account, we propose that the cAMP molecule binds Xepac protein enabling it to activate the IP3/Ca2+ pathway, which is necessary to start and maintain X. laevis vitellogenin uptake.
Subject(s)
Calcium/metabolism , Inositol 1,4,5-Trisphosphate/metabolism , Oocytes/physiology , Oogenesis/physiology , Vitellogenins/metabolism , Xenopus Proteins/metabolism , Animals , Calcimycin/pharmacology , Calcium Ionophores/pharmacology , Calcium Signaling/drug effects , Cyclic AMP/analogs & derivatives , Cyclic AMP/metabolism , Cyclic AMP/pharmacology , Female , Gene Expression Regulation , Oocytes/drug effects , Oogenesis/drug effects , Signal Transduction , Thionucleotides/pharmacology , Xenopus Proteins/genetics , Xenopus laevisABSTRACT
The aim of the present study was to show the participation and physiological role of calmodulin (CaM) and cAMP during vitellogenin endocytic uptake in the amphibian Xenopus laevis. The results showed a differential distribution of CaM in the ovary follicles during oogenesis. The CaM intracellular localization was not affected by gap junction's downregulation and CaM inhibition did not completely abolished the endocytic activity of oocytes. We showed that cAMP was able to completely rescue the endocytic competence in follicles in which gap junctional communication had been disrupted by octanol. Moreover cAMP was capable of restoring oocyte endocytic capability in the presence of octanol and stelazine, a CaM inhibitor. We propose that, in Vtg uptake regulation, cAMP is upstream of CaM during the endocytic signalling pathway.