Nanoemulsion of Spiramycin against Tachyzoites of Toxoplasma gondii, RH Strain: Preparation, Toxicology, and Efficacy Studies.

Background: Toxoplasma infection is caused by Toxoplasma gondii, which is an intracellular protozoan parasite. This infection consequently lead various congenital disabilities during pregnancy in patients. Spiramycin (Spi), a macrolide antibiotic, is typically recommended for T. gondii infection in pregnant women. We aimed to prepare the nanoemulsion of spiramycin (NE-Spi) and to evaluate the activity of this formulation in tachyzoites of T. gondii, RH strain. Methods: This study was conducted in 2019-2021 at the School of Public Health, Tehran University of Medical Sciences, Tehran, Iran. NE-Spi was prepared by spontaneous emulsification. The effects of this nanoemulsion on the viability of cultured cells were measured using MTT assay. To estimate the effects of NE-Spi on tachyzoites of T. gondii, RH strain, different concentrations of NE-Spi, S-Spi (suspension of spiramycin), and NE (nanoemulsion without any spiramycin) were added to tachyzoites and then stored for 30, 60, 90, 120 min and 24 h in 250 µg/ml concentration at room temperature. Finally, Tachyzoites mortality rates were evaluated by trypan blue staining. Of note, flow cytometry was conducted to confirm the obtained results. Results: The final particle size of NE-Spi was calculated to be 11.3 nm by DLS and TEM. Thereafter, using MTT assay, in 62.5 µg/ml concentration of NE-Spi, the Vero cells viability was obtained as 82%. The highest mortality rates of tachyzoites of T.gondii, RH strain were observed at 250 µg/ml concentration and after 120 min of exposure, but it was not significantly different from 24 h of exposure. Conclusion: NE-Spi has lethal efficacy on T. gondii RH strain in-vitro.

Protoscolicidal effects of curcumin nanoemulsion against protoscoleces of Echinococcus granulosus.

BACKGROUND: The aim of the present study was to assess in vitro protoscolicidal effects of curcumin nanoemulsion (CUR-NE) against protoscoleces of cystic echinococcosis (CE)/hydatid cysts. METHODS: The CUR-NE was prepared via spontaneous emulsification of soybean as the oil phase, a mixture of Tween 80 and Tween 85 as the surfactant, ethanol as the co-surfactant and distilled water. Various concentrations of CUR-NE (156, 312, 625 and 1250 µg/ml) were exposed to collected protoscoleces of infected sheep liver hydatid cysts for 10, 20, 30, 60 and 120 min. Viability of the protoscoleces were assessed using eosin exclusion test. Morphological changes of the protoscoleces were observed using differential interference contrast (DIC) microscopy. RESULTS: The mean particle size and zeta potential of CUR-NE included 60.4 ± 14.8 nm and - 16.1 ± 1.1 mV, respectively. Results showed that the viability of the protoscoleces decreased significantly with increases in CUR-NE concentrations (p < 0.001). The mortality rates of protoscoleces with exposure to concentrations of 1250 and 625 µg/ml of CUR-NE for 60 min were 94 and 73.33%, respectively. Mortality of the protoscoleces was 100% after 120 min of exposure to 1250 and 625 µg/ml concentrations of CUR-NE. Using NIC microscopy, extensively altered tegumental surface protoscoleces was observed after protoscoleces exposure to CUR-NE. CONCLUSION: The findings of the present study revealed the in vitro protoscolicidal potential of CUR-NE. Therefore, CUR-NEs are addressed as novel protoscolicidal agents, which can be used as an alternative natural medicine to kill the protoscoleces, owing to their low toxicity and significant inhibition potency. However, further studies are necessary to investigate pharmacologic and pharmacokinetics of CUR-NEs.

Assessment of an In-House Enzyme-Linked Immunosorbent Assay and IgG Avidity Test Based on SAG1 and GRA7 Proteins for Discriminating between Acute and Chronic Toxoplasmosis in Humans.

To improve serodiagnostic methods for diagnosis of acute from chronic toxoplasmosis, an economical in-house enzyme-linked immunosorbent assay (ELISA) for measuring Toxoplasma-specific IgG, IgM, and IgG avidity has been developed and assessed based on use of various Toxoplasma gondii antigens, including SAG1, GRA7, and a combination of SAG1 and GRA7 (SAG1+GRA7), as well as Toxoplasma lysate antigens (TLAs). Performances of in-house IgM, IgG, and IgG avidity assays were compared to those of ELISA commercial kits and VIDAS Toxo IgG avidity. A set of 138 sera from patients with acquired T. gondii infection and seronegative people were assessed. Receiver operating characteristic (ROC) analysis revealed an area under curve (AUC) of 0.98, 0.97, 0.99, and 0.99 for IgM-TLAs, IgM-SAG1, IgM-GRA7, and IgM-SAG1+GRA7, respectively. Furthermore, AUC was calculated as 0.99, 0.99, 0.98, and 0.99 for IgG-TLAs, IgG-SAG1, IgG-GRA7, and IgG-SAG1+GRA7, respectively. The current study showed that GRA7 included 100% sensitivity for the detection of Toxo IgM, while SAG1 included 89.7% sensitivity. Furthermore, the highest specificity (97.2%) to detect Toxo IgM was achieved using SAG1+GRA7 antigen. For the detection of Toxo IgG, the highest sensitivity (100%) was recorded for SAG1+GRA7, followed by TLAs (97.9%). The SAG1+GRA7 showed the greatest potential for assessing avidity of IgG antibodies, with 97.1% sensitivity and 96.6% specificity compared to those of VIDAS Toxo IgG avidity. The preliminary results have promised better discriminations between acute and chronic infections using a combination of SAG1 and GRA7 recombinant antigens compared to those using TLAs.

Unravelling Toxoplasma treatment: conventional drugs toward nanomedicine.

Toxoplasma gondii is a worldwide protozoan parasite that infects almost all warm-blooded animals. Although human toxoplasmosis is mostly latent, pregnant women and immunocompromised patients need effective treatment. There are drugs of choice for treatment of toxoplasmosis; however, due to their side effects and/or their disease stage-specificity, prescription of them is limited. During recent years, nanomedicine has been employed to overcome limitations of conventional drugs. Here, we provided a state-of-the-art review of experimental toxoplasmosis treatment using nanotechnology.

Molecular Characterization of Fasciola spp. from Some Parts of Iran.

BACKGROUND: Identification of liver flukes, Fasciola hepatica, and Fasciola gigantica by morphometric parameters is not always reliable due to the overlapping measurements. This study aimed to characterize the liver flukes of animals from different parts of Iran by the genetic markers, ITS1, and COXI. METHODS: We collected flukes from infected livestock in six provinces of Iran from Sep to Nov 2016. The flukes were identified by amplification of a 680 bp sequence of ITS1 locus followed by a restriction fragment polymorphism (RFLP) assay. The genetic diversity among isolates was evaluated by amplification and sequencing of a 493 bp fragment of the COXI gene. RESULTS: We obtained 38 specimens from Khuzestan, 22 from Tehran, 10 from Isfahan, 10 from Mazandaran, 4 from Kurdistan, and 3 from Ardabil provinces. PCR-RFLP analysis revealed two patterns, representing F. hepatica, and F. gigantica. Fifty specimens from cattle and sheep exhibited F. hepatica pattern and 37 from the cattle, sheep, buffalo, and goat that of F. gigantica. The phylogeny based on COXI revealed two distinct clades separating F. hepatica from F. gigantica. In our phylogeny, the Iranian F. gigantica isolates showed a distinct separation from the African flukes, while grouped with the East Asia specimens demonstrating a common ancestor. The F. hepatica isolates clustered with the flukes from different parts of the world, including East Asia, Europe, and South America. CONCLUSION: The present study revealed a substantial genetic difference between F. gigantica populations of Asia and Africa, while F. hepatica isolates from different parts of the world shared high similarities.