Evaluation of synergistic therapeutic effect of shark cartilage extract with artemisinin and glucantime on visceral leishmaniasis in BALB/c mice

Objective(s): Because leishmaniasis is related to the impaired functioning of T-cells, the use of an immunomodulator can increase the efficacy of antileishmanial therapy in visceral leishmaniasis. In this study, we used shark cartilage extract with artemisinin and glucantime against visceral leishmaniasis in BALB/c mice, and evaluated the synergistic therapeutic effect. Materials and Methods: The culturing method and quantitative real-time PCR by using the kDNA gene was used to detect parasite loads in the spleen and liver. INF-γ and IL-4 cytokine levels and survival rates were assayed.Results: The drug therapy with target drugs reduced parasite burden in the spleen and liver significantly. Although parasite burden was lower in the artemisinin treated group than in the glucantime treated group (

Molecular Identification and Intra-species Variations among Leishmania infantum Isolated from Human and Canine Visceral Leishmaniasis in Iran

Background: In Iran, both forms of cutaneous (CL) and visceral leishmaniasis (VL) have been re-ported; so the accurate species identification of the parasite(s) and the analysis of genetic diversity are necessary. Methods: The investigation was conducted from 2014 to 2015 in the northwest and south of Iran, where VL is endemic (7 provinces). Blood samples of patients and infected dogs were collected and sera separated for serologic examinations (DAT, rK39). Spleen or bone marrow samples from infected dogs were also collected to confirm the infection. DNAs of 70 samples amplified by targeting a partial sequence of ITS (18S rRNA–ITS1–5.8S rRNA–ITS2) gene. All the amplicons were sequenced and analyzed with restriction fragment length polymorphism (RFLP) using the TaqI enzyme. Results: The cause of all 70 VL cases, were L. infantum, so, the dominant specie is L. infantum. The sequencing results of all VL cases and RFLP analysis corroborate each other. Discrimination of Iranian Leishmania isolates using ITS gene gives us this opportunity to detect, identify and construct the phylogenetic relationship of Iranian isolates. In addition, detection and differentiation of Leishmania spp. DNA was confirmed by amplification of variable area of the minicircle kDNA (conserved sequence blocks (CSB)). Conclusion: Low divergence and high likelihood were seen among L. infantum isolates of human and dogs from Iran with a very slight divergence was seen between isolates from northwest and south of Iran, thus grouped in a unique clad. No correlation was observed between intraspecies divergence and geographic distribution of the isolates

Prevalence and Risk Factors of Toxoplasma gondii Infection among Pregnant Women in Hormozgan Province, South of Iran

Background: Toxoplasmosis can cause miscarriage or complications in the fetus. Diagnosis and treatment of this disease by anti-parasitic drugs especially in early pregnancy can help to prevent fetal infection and its complications. This study aimed to determine T. gondii infection in pregnant women, evaluate risk factors in the transmission of the disease and congenital toxoplasmosis. Methods: Overall, 360 sera of pregnant women from 5 cities in the Hormozgan Province in southern Iran with different climate were evaluated from 2015-2016 for T. gondii infection by using ELISA method and positive cases of IgM and IgG were tested again using Avidity IgG ELISA. All cases were evaluated according to climate, acute and chronic of toxoplasmosis, number of pregnancy and abortion, epidemiological factors and food habits. Results: Among 360 specimens T. gondii IgG + IgM antibodies were found positive in 0. 8% subjects and also 27% of samples had IgG seropositivity. A significant relationship was observed between age, sampling place, consumption of raw and half cooked meat, history of contact with cats, abortion history, number of children, and parity with IgG positive. In Avidity IgG ELISA test, 13 people with low avidity, 3 people with borderline avidity were reported. Conclusion: 72. 2% of the population had no antibody against the disease that this could be a warning to the people and requires education of preventive and prenatal care and routine screening of women at childbearing age

Anti-leishmanial activities of selenium nanoparticles and selenium dioxide on Leishmania infantum

Leishmania infantum is one of the important causes of visceral leishmaniasis in many countries. There are different complications for treatment of leishmaniasis such as toxicity and drug resistant. So far, there isn’t any information about the effects of selenium nanoparticles and selenium dioxide (chemical form of selenium) on Leishmania parasites; hence, the aim of the present study is to investigate in vitro effects of six dilutions of these drugs on L. infantum. Anti-leishmanial activities were studied by adding different dilutions of 2.5, 5, 10, 25, 50, and 100 μg/ml of the drugs into promastigote cultures. Promastigote cytotoxicity was tested using the colorimetric MTT assay. Anti-amastigote activity was assessed in peritoneal macrophages of BALB/c mice. Also, cytotoxic effect of these drugs was evaluated on uninfected macrophages. The results showed that both of drugs have dose-dependent anti-leishmanial activities. Selenium NPs have more growth-inhibitory effect on promastigotes than SeO2; while the IC50 (50 % inhibitory concentration) was determined to be 25 and 50 μg/ml, respectively. The mean numbers of amastigotes per macrophage in selenium NPs-treated groups were less than SeO2-treated and control groups. The IC50 of selenium NPs was 10 μg/ml and SeO2 was 25 μg/ml for amastigotes. Also, the IC50 of selenium NPs and SeO2 for uninfected macrophages were calculated to be 100 and 50 μg/ml, respectively. In addition, selenium NPs has less cytotoxic effect than SeO2 on uninfected macrophages. These findings suggest that selenium NPs have more anti-leishmanial properties and less cytotoxic effects than SeO2 against L. infantum

In Vitro Antiparasitic and Apoptotic Effects of Antimony Sulfide Nanoparticles on Leishmania infantum

Visceral leishmaniasis is one of the most important sever diseases in tropical and subtropical countries. In the present study the effects of antimony sulfide nanoparticles on Leishmania infantum in vitro were evaluated. Antimony sulfide NPs (Sb2S5) were synthesized by biologicalmethod fromSerratia marcescens bacteria. Then the cytotoxicity effects of different concentrations (5, 10, 25, 50, and 100 μg/mL) of this nanoparticle were assessed on promastigote and amastigote stages of L. infantum. MTTmethodwas used for verification results of promastigote assay. Finally, the percentages of apoptotic, necrotic, and viable cells were determined by flow cytometry. The results indicated the positive effectiveness of antimony sulfide NPs on proliferation of promastigote form. The IC50 (50% inhibitory concentration) of antimony sulfide NPs on promastigotes was calculated 50 μg/mL. The cytotoxicity effect was dose-dependent means by increasing the concentration of antimony sulfide NPs, the cytotoxicity curve was raised and the viability curve of the parasite dropped simultaneously. Moreover, the IC50 of antimony sulfide NPs on amastigote stage was calculated 25 μg/mL. On the other hand, however, antimony sulfide NPs have a low cytotoxicity effect on uninfected macrophages but it can induce apoptosis in promastigote stage at 3 of 4 concentrations

Molecular and Morphological Data Confirmed First Record of Abbreviata kazakhstanica Markov and Paraskiv, 1956 (Spirurida: Physalopteridea) in Iran

Background: The genus Abbreviata (Spirurida: Physalopteridea) currently contains 47 species. Physalopteridae nematodes infect a large number of vertebrates, including mammals, birds, reptiles and amphibians. The current study is a report of the first morphological and molecular identification of A. kazakhstanica (Spirurida: Physalopteridea) in Pseudopus apodus in Iran. Methods: Eleven road-killed P. apodus, were collected from, Iran during 2016-2018. The nematodes were isolated from stomach. After morphological study, the genomic DNA of the parasites was extracted using CTAB method. The DNA was used for PCR amplification of cytochrome c oxidase subunit I (cox1). The PCR products were sequenced, the sequence data were analyzed and multiple alignments were conducted using the Clustal Omega. Results: After detailed microscopic examination, the A. kazakhstanica was identified. The cox1 sequences confirmed the species of helminth. The new sequences of A. kazakhstanica were submitted to GenBank under the accession number MK578751-2. Conclusion: Regarding the limited data on parasitological status of Iranian reptiles, more specific and comprehensive investigations are needed to identify the parasitic fauna Keywords Abbreviata kazakhstanica, Molecular identification, Pseudopus apodus, Physalopteridea, Ira

Molecular Identification and Phylogenetic Classification of Leishmania spp. Isolated from Human Cutaneous Leishmaniasis in Iran: A Cross-sectional Study

Background: In Iran, both forms of cutaneous (CL) and visceral leishmaniasis (VL) have been reported; so the accurate species identification of the parasite(s) and the analysis of genetic diversity are necessary. Methods: The smears were collected from lesions samples of 654 patients with CL, who attended local health centers in 12 provinces of Iran during 2013-2015. The smears were checked for the presence of amastigotes by light microscopy. DNA of 648 Leishmania isolates, amplified by targeting a partial sequence of ITS (18S rRNA–ITS1–5.8S rRNA–ITS2) gene. Twenty-five of all the amplicons were sequenced and analyzed with restriction fragment length polymorphism (RFLP) using the Taq1 enzyme. Results: All the smears were positive microscopically. The PCR-RFLP analysis revealed that 176 (27%) CL patients were infected with L. tropica and, 478 (73%) with L. major. The dominant species in all over Iran is L. major. The sequencing results of all CL patients and RFLP analysis confirmed each other. Based on our phylogenetic tree, 25 ITS DNA sequences were grouped into two clusters representing L. major and L. tropica species. Phylogenetic tree derived from the ITS sequences supports a clear divergence between L. major from the other species. Conclusion: Discrimination of Iranian Leishmania isolates using ITS gene gives us this opportunity to detect, identify, and construct the phylogenetic relationship of Iranian isolates

In vitro and in vivo antileishmanial effects of aloe-emodin on Leishmania major

Cutaneous leishmaniasis is a common parasitic disease that is endemic in some parts of Iran. The drugs of choice used for leishmaniasis therapy are associated with a risk of recurrence and serious adverse effects. Therefore, finding a safe and effective treatment is of great importance. In the present study, the effect of aloe-emodin on the growth of Leishmania major amastigotes was evaluated under in vitro conditions. In addition, the efficacy of a topical of aloe-emodin ointment was investigated in BALB/c mice with cutaneous leishmanial ulcers. Different concentrations (40 μg/mL, 80 μg/mL, 120 μg/mL, and 160 μg/mL) of aloe-emodin were tested on Leishmania amastigotes twice: 24 hours and 48 hours. The induced apoptosis and necrotic effects of two concentrations (40 μg/mL and 120 μg/mL) of aloe-emodin on promastigotes were investigated by flow cytometry. Under the in vivo condition, aloe-emodin ointment efficacy was evaluated at two concentrations (i.e., 0.1% and 1%). Serum indicator factors of the test and control groups were tested to evaluate the toxic effects of this compound on the liver and kidney. Results showed that aloe-emodin inhibited the growth of Leishmania amastigotes and induced apoptosis in promastigotes. Topical application of aloe-emodin ointment likewise reduced the ulcer size. No significant differences in biochemical analysis were observed between the control and treated groups. In conclusion, aloe-emodin showed antileishmanial effects under in vitro and in vivo conditions and may be used in clinical trials

Association of Toxoplasma Gondii Infection With Diabetes Mellitus Using Nested-PCR and Sequencing

Background: Toxoplasmosis is a global public health concern with severe complications, particularly in pregnant women and immunosuppressed patients. Herein, we revealed the first genotypic evidence of Toxoplasma gondii (T. gondii) in diabetic and non-diabetic individuals referred to Imam Sajjad Hospital, Tehran, Iran, using nested-Polymerase Chain Reaction (PCR) and sequencing. Materials and Methods: We collected 98, 95, and 94 blood samples from Type-1 Diabetes Mellitus (T1DM), Type-2 Diabetes Mellitus (T2DM), and control subjects from February to August 2018, and their DNA was extracted. After amplification and visualization of dense granule antigen 6 gene (344 bp) using external and internal primer pairs, the positive samples were sent for sequencing. Results: Among 287 individuals, three T1DM patients (two IgG, one IgM) and one T2DM patient (IgG) were found positive by molecular method, which proved to be type III strain by sequencing. Conclusion: To the best of our knowledge, this is the first genotyping investigation on T. gondii in diabetic individuals. Further research should be done to better realize the association between Toxoplasma genotypes and the outcome of diabetes mellitus