In vitro therapeutic effect of Hemiscorpius lepturus venom on tachyzoites of Toxoplasma gondii.

Pyrimethamine which is a main anti-Toxoplasma gondii drug has a serious side and toxic effects on the host. Accordingly, the development of new treatment options for toxoplasmosis with less toxic effects, low teratogenicity and parasiticidal effect against the various stage of T. gondii are dramatically crucial. Currently, natural molecules from scorpion and snake venoms are widely used as an alternative treatment against human disease, these compounds considered to be safe and to have low toxicity in comparison with synthetic drugs. Therefore, the goal of our study was to investigate the anti-Toxoplasma gondii activities of Hemiscorpius lepturus venom. We measured cytotoxicity of H. lepturus whole venom on Vero cells as well as effectiveness of this compound on viability of T. gondii applying colorimetric assay, according to mitochondrial oxidation of the MTT reagent (Methylthiazol tetrazolium 98%). The results of this study indicated that the H. lepturus whole venom has an anti-Toxoplasma effects with less toxic effect on Vero cells. Also, the T. gondii tachyzoites were treated with H. lepturus venom reached better results in comparison with Pyrimethamine-treated group. This research will serve as a base for future studies on toxoplasmosis and suggest a role for scorpion venom in promoting natural drugs.

Identification and Determination of the Geographical Distribution of Freshwater Snails in Lorestan, Iran.

Snails are creatures present in various ecosystems that, in addition to being present in human surroundings, some of them are also important in veterinary medicine and medicine as the intermediate hosts of Digenean trematodes. The present study was conducted to identify and determine the geographical distribution of freshwater snails and investigate the relationship of variables, such as season and geographical region, with snail species and dispersion in Lorestan in the west of Iran. A total of 4400 samples of freshwater snails were collected using the multistage sampling method (i.e., stratified, cluster, and randomized) from 110 points in five geographical regions in four seasons and then identified based on their morphological characteristics by diagnostic keys. The ArcGIS software (version 10.3) was used to evaluate the spatial distribution of the freshwater snails. In this study, seven species of freshwater snails were identified in six families belonging to six genera, namely Melanopsis doriae (6.30% of the variation in species), Theodoxus doriae (5.55%), Bithynia tentaculata (43.22%, the dominant species), Physa acuta (24.98%), Lymnaea truncatula (9.75%), Gyraulus euphraticus (8.18%), and Lymnaea gedrosiana (2.02%). The geographic distribution of freshwater snails was recorded across five regions in 22 points per region for every season. The spatial distribution maps showed that the distribution of freshwater snails varies according to region and season (P<0.001). The obtained results revealed the effects of season and geographical region on the distribution and population density of snails in the province. These data can be used for the implementation of control programs against parasitic diseases in the region, including trematodes.

Cloning and Expression of Immunogenic Regions of EMA-1 Gene of Theileria equi From Infected Horses.

Diversity among the pathogenic strains of Theileria equi (T. equi), a major agent of equine piroplasmosis, can affect the appropriate detection of parasite and host immunization. Production of recombinant surface proteins from an infected horse in natural endemic area provides a reliable tool for immunodiagnosis of parasite. Regarding this, the present study was targeted toward the cloning, expression, and purification of the immunogenic regions of equine merozoite antigen 1 (EMA-1 gene), as one of the most important immunodominant surface proteins in T. equi, from naturally infected horses in Iran. The immunogenic region of EMA-1 gene was amplified using the blood of infected horses. EMA-1 gene was cloned into pET26b vector. Then, recombinant plasmids (pET 26b-EMA-1) were transformed into competent E. coli BL21 (DE3) cells. Cloning was confirmed by polymerase chain reaction (PCR), restriction enzyme assays, and DNA sequence analysis. The recombinant protein was expressed using isopropyl β-D-1-thiogalactopyranoside as an inducer, purified using nickle-nitrilotriacetic acid column, and then confirmed by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and dot blot analysis utilizing Anti-His Tag antibody. Furthermore, the immunoreactivity of recombinant protein against the serum of the infected horses was evaluated using dot blot analysis. The PCR product analysis showed a 750-bp band belonging to immunogenic regions of EMA-1 gene. Sequence analysis revealed that cloned EMA-1 and protein had 94% and 97% homology to EMA-1 sequences submitted to GenBank from different countries, respectively. Restriction enzyme and sequence analyses confirmed the subcloning and correction of the orientation of inserted gene. The SDS-PAGE analysis confirmed the expression of EMA-1 protein with a 28-kDa band. The results of the dot blot analysis revealed that the horse serum containing antibody against T. equi could react with the purified recombinant protein. Purified EMA-1 protein can be used as a reliable tool for the future development of diagnostic tests or vaccines.

Detection of specific hydatid antigens and antibodies in serum and urine of experimentally infected sheep.

Immunodiagnostic confirmation of cystic human hydatidosis is frequently required before surgical intervention or of chemotherapy. However, it remains inadequate to detect specific antibodies or antigens in some confirmed cases of echinococcosis. This study was carried out to investigate the accuracy of three different immunodiagnostic tests for detection of specific circulating antigens or antibodies in the serum and urine of 13 experimentally infected sheep. For this purpose, Echinococcus granulosus were collected from small intestine of experimentally infected dogs, and 2000 taenid eggs were orally administered to each of the 13 sheep. There were six other sheep, which were kept as the control group. Biweekly serum and urine samples were collected from all the sheep for 4 months after infection. The sera were subjected to indirect hemagglutination test and the concentrated urine samples were subjected to coagglutination and counter immunoelectrophoresis tests. The results revealed that the sensitivity of these tests in detecting the hydatid antigens in the urine or antihydatid antibodies in the serum of the infected sheep reached their maximum in 12th and 13th week after infection; then it decreased in the following weeks. Examination of the non-infected sheep samples throughout the experiment showed that the aforesaid findings were specific only to the infected sheep. It seems that the appearance of specific hydatid antigen in urine and its antibodies in the serum were simultaneous. Although these tests are highly specific, false negative outcomes were encountered in their detection of cystic echinococcosis. In general, it seems rational to establish some series of diagnostic procedures in order to reveal antibodies and antigen of metacestode in serum and urine of the patients.