Promising effects of 1,8 Cineole to control Giardia lamblia infection: Targeting the inflammation, oxidative stress, and infectivity.

Reportedly, synthetic drugs such as metronidazole, furazolidone, tinidazole, and quinacrine are used for the treatment of giardiasis but are associated with adverse effects. In this study, we aimed to investigate the in vitro and in vivo effects of eucalyptol (ECT, 1,8 cineole) alone and in combination with metronidazole (MNZ) on Giardia lamblia. The effects of ECT on cell viability, plasma membrane permeability, and gene expression levels of adenylate cyclase (AK) and extracellular signal kinases 1 and 2 (ERK1 and ERK2) in trophozoites of G. lamblia were assessed. In vivo, the effects of ECT alone and in combination with MNZ were assessed on mice infected with G. lamblia. In addition, the gene expression of inflammatory genes (e.g., TNF-α, IL-1ß, and IL-10) and antioxidant genes (catalase (CAT), superoxide dismutase 1 (SOD1), glutathione peroxidase 2 (GPX2)) was determined by real-time PCR. The IC50 values of ECT, MNZ, and ECT+MNZ on trophozoites were 30.2 µg/mL, 21.6 µg/mL, and 8.5 µg/mL, respectively. The estimated Fractional inhibitory concentration index (FICI) values for ECT and MNZ were 0.28 and 0.39, respectively. The application of ECT on G. lamblia trophozoites resulted in a dose-dependent increase in plasma membrane permeability, particularly at concentrations of ½ IC50 and IC50 (P < 0.05). The treatment of infected mice with various doses of ECT, mainly in combination with MNZ for 7 days, resulted in a significant decrease (P < 0.001) in the average number and viability of cysts. ECT, especially when combined with MNZ, caused a significant (P < 0.001) reduction in the expression of TNF-α and IL-6 genes, and an increase (P < 0.05) in the expression of IL-10 genes. ECT alone and mainly in combination with MNZ leads to a significant (P < 0.001) increase in the gene expression of CAT, SOD, and GPX genes. These findings demonstrate that the use of ECT in these doses, even for 14 days, does not have any toxic effects on the function of vital liver and kidney tissues. The study findings confirmed the promising effects of ECT against G. lamblia infection both in vitro and in vivo. Considering the possible mechanisms, ECT increases plasma membrane permeability and reduces the expression levels of infectivity-related genes. In addition, ECT suppresses inflammation and oxidative stress, controlling giardiasis in mice. More studies are needed to clarify these findings.

Molecular Epidemiology and Associated Risk Factors of Parasites in Oral Cavity of Children with Malignancies in Western Iran.

Background: This survey was designed to study the molecular epidemiology and risk factors of Entamoeba gingivalis and Trichomonas tenax in children with underlying malignancies and those on chemotherapy in Lorestan province, West of Iran. Methods: The present cross-sectional descriptive study was performed on children who suffering from different types of malignancies or receiving treatment by chemotherapy referring to oncology section of hospitals of Lorestan Province, Iran during May 2021 to April 2022. The frequency of oral cavity protozoa was investigated using microscopic and conventional polymerase chain reaction (PCR). Results: E. gingivalis and T. tenax parasites were found in 23 (25.5%) by microscopic method and 28 (31.1%) using PCR in children with malignancy. Among positive samples, 20 (71.4%) were infected with E. gingivalis; whereas 8 (28.6%) of the participants were positive for T. tenax. In the multivariate model, living in rural regions (OR= 3.437; 95% CI= 1.22-9.63; p=0.019) and using mouthwash (OR= 0.082; 95% CI= 0.018-0.37; p<0.001) were significantly related with the frequency of oral cavity parasites. Conclusion: Our results showed the high frequency of oral cavity parasites in children who suffering malignancies or receiving treatment by chemotherapy in Lorestan province, Iran. The awareness of the main risk factors for oral cavity parasites particularly using mouthwash is necessary in improving public and oral health strategies in children with cancer. Consequently, oncologist and dental practitioners must be aware to identify and manage oral health concerns in in children who suffering from different types of malignancies to prevent the oral diseases and infections.

Prevalence of Toxoplasma gondii in HIV-Infected Individuals in Iran: A Systematic Review and Meta-Analysis.

INTRODUCTION: Toxoplasma gondii is an obligate intracellular protozoan that can infect almost all warm-blooded animals, including humans. Patients with co-infection with toxoplasmosis and HIV have a 30-40% risk of developing toxoplasmosis encephalitis. This study aimed to describe the epidemiology and burden of Toxoplasma gondii in HIV-infected individuals in Iran. METHODS: We searched the five English databases (Science Direct, PubMed, Scopus, Ovid, Embase, and Cochrane) and four Persian databases (Scientific Information Database (SID), Iran Medex, Iran Doc, and Magiran) with the terms of (Toxoplasma gondii OR "toxoplasmosis") AND (HIV OR "AIDS" OR immunodeficiency OR acquired immune deficiency syndrome) AND (Seroprevalence) AND (Seroepidemiologic Studies) AND (Elisa OR IgG) AND (PCR) AND (Iran) by two authors up to Feb 2021. Studies were included if they investigated people with HIV infection and presented data that allowed us to establish the prevalence of Toxoplasma gondii infection in Iran. RESULTS: According to the inclusion/exclusion criteria, 15 studies were selected. A total number of 2275 HIV-infected individuals were tested and evaluated for toxoplasmosis from 2005 up to 2018 in different regions of Iran. The weighted overall prevalence of toxoplasmosis in HIV-infected individuals with Elisa was obtained using a random-effects model, which was estimated at 47% (95% CI = 31% - 62%). Also, the Weighted overall prevalence of toxoplasmosis in HIV-infected individuals with PCR was obtained using a random-effects model, which was estimated at 7% (95% CI = 3% - 12%). CONCLUSION: According to the results of this study, it can be clearly understood that a large population of HIV patients living in Iran have toxoplasmosis. Therefore, due to the high susceptibility of these groups to toxoplasmosis, healthcare professionals must consider measures such as training in the ways of transmission and prevention of the infection to this high-risk group in order to reduce the risk of infection.

Nanoparticles as Potent Agents for Treatment of Schistosoma Infections: A Systematic Review.

Background: Schistosomiasis is an acute and chronic parasitic disease caused by blood flukes of the genus Schistosoma. The current drugs for treating schistosomiasis are associated with some side effects. Objective: The aim of this systematic study was an overview of the treatment of diseases caused by Schistosoma based on nanoparticles. Methods: In the present systematic research with keywords "Schistosoma", "parasitism", "anti-Schistosoma activity", "nanoparticles", "metal nanoparticles", "silver nanoparticles", "gold nanoparticles", "polymer nanoparticles", "PLGA nanoparticles", "nanoemulsions", "in vitro", and "in vivo" from five English-language databases, including ScienceDirect, europePMC, PubMed, Scopus, Ovid, and Cochrane were searched from 2000 to 2022 by 2 researchers. Results: In the initial search, 250 studies were selected. Based on the inclusion and exclusion criteria, 27 articles were finally selected after removing duplicate, unrelated, and articles containing full text. In present article, the most nanoparticles used against Schistosoma were gold nanoparticles (22%). Conclusions: The results indicate the high potential of various nanoparticles, including metal nanoparticles, against Schistosoma. Also, the remarkable anti-schistosomal activity of nanoparticles suggests their use in different fields to eliminate this pathogenic microorganism so that it can be used as an effective candidate in the preparation of anti-schistosomal compounds because these compounds have fewer side effects than chemical drugs. Ther Res Clin Exp. 2023; XX:XXX-XXX).

Immune-enhancing activity of Astragalus maximus extract for inhibiting the Toxoplasma gondii infection: experimental research.

Background: This study aimed to evaluate the in-vitro anti-Toxoplasma effects and cytotoxicity effects of Astragalus maximus chloroformic extract (AMCE) on the T. gondii Rh strain. Methods: In-vitro effects of AMCE (2-64 µg/ml) on tachyzoites were measured by MTT assay for 48 h. The effects of AMCE on infectivity rate and intracellular parasites into macrophage cells (J774-A1) were evaluated. The Griess reaction assay and quantitative real-time PCR were used to determine the nitric oxide (NO) and the mRNA expression levels of IFN-γ and iNOS in infected J774-A1 macrophage cells. Results: The mortality rate of the parasites significantly (P<0.001) increased in a dose-dependent manner with an IC50 value of 9.85 µg/ml. The rate of infection and the mean number of intracellular tachyzoites in macrophage cells were significantly reduced (P<0.001) after exposure of the macrophage cells to AMCE. The mRNA expression levels of IFN-γ, iNOS, and NO production in macrophage cells after treatment with the AMCE were increased, especially at the concentration of ½ IC50 and IC50 (P<0.001). Conclusion: The findings of the current in vitro investigation revealed favorable anti-Toxoplasma effects of AMCE against tachyzoites and intracellular forms of T. gondii. Despite the fact that the accurate anti-Toxoplasma mechanisms of AMCE are not clear, our results showed that triggering NO production and cellular immunity can be considered as the main mechanisms of action of AMCE for controlling and eliminating T. gondii. However, further surveys are mandatory to assess the efficacy and safety of AMCE in an animal model and its accurate mechanisms of action.

Synthesis, characterization, and antiparasitic effects of zinc oxide nanoparticles-eugenol nanosuspension against Toxoplasma gondii infection.

Background: In this study, zinc oxide nanoparticles-coated with eugenol (ZnO@Eug) were synthesized and evaluated as a nanosuspension (NSus) formulation against Toxoplasma gondii in vitro and in vivo. Methods: An anti-Toxoplasma activity assay for ZnO@Eug NSus was conducted in vitro, ex vivo, and in vivo. FTIR spectroscopy confirmed the formation of ZnO@Eug NSus by detecting several functional groups involved; EDX and SEM demonstrated the grain of ZnO-NPs embedded with Eug and compositional purity. Results: Surface charge (ZP) and size distribution (DLS) of ZnO@Eug NSus were determined to be -22.7 mV and 109.6 nm, respectively. According to the release kinetics, approximately 60% of the ZnO-NPs and Eug were released in the first 45 min. In the cytotoxicity assay, ZnO-NPs, Eug, and ZnO@Eug NSus had IC50 values of 71.85, 22.39, and 2.02 mg/mL, respectively. The therapeutic efficacy of ZnO@Eug against T. gondii was 56.3%, which was not significantly different from that of spiramycin (58.9%) (Positive-control). The tissue tachyzoites in the liver, spleen, and peritoneum were less than 50% in groups treated with Eug, spiramycin, and ZnO@Eug NSus compared to the control. ZnO@Eug-treated groups showed a survival rate of up to 13 days. Conclusions: The ZnO@Eug NSus demonstrated antiparasitic activity against T. gondii with minimal toxic effects and high efficiency in increasing the survival of infected mice. The nanoformulations of ZnO-NPs incorporated with Eug could, in the future, be considered for treating toxoplasmosis in humans and animals if a detailed study was conducted to determine the precise dose and measure side effects.

Antileishmanial, cellular mechanisms, and cytotoxic effects of green synthesized zinc nanoparticles alone and in combined with glucantime against Leishmania major infection.

BACKGROUND: We decided to investigate the antileishmanial, cellular mechanisms, and cytotoxic effects of green synthesized Zinc nanoparticles (ZnNPs) alone and combined with glucantime against Leishmania major infection. METHODS: The effect of green synthesized ZnNP on L. major amastigote was studied through macrophage cells. The mRNA expression level of iNOS and IFN-γ followed by the exposure of J774-A1 macrophage cells to ZnNPs was assessed by Real-time PCR. The Caspase-3-like activity of promastigotes exposed to ZnNPs was studied. Effects of ZnNPs alone and combined with glucantime (MA) were studied on cutaneous leishmaniasis in BALB/c mice. RESULTS: ZnNPs displayed the spherical shape with sizes ranging from 30 to 80 nm. The obtained IC50 values for ZnNPs, MA, and ZnNPs + MA were 43.2, 26.3, and 12.6 µg/mL, respectively; indicating the synergistic effects of ZnNPs in combination with MA. CL lesions had completely improved in the mice received with ZnNPs in combination with MA. The mRNA expression level of iNOS, TNF-α, and IFN-γ was dose-dependently (p < 0.01) upregulated; whereas it was downregulated in IL-10. ZnNPs markedly stimulated the caspase-3 activation with no significant toxicity on normal cells. CONCLUSION: Based on these in vitro and in vivo results, green synthesized ZnNPs, mainly along with MA, showed that has the potential to be introduced as a new drug for CL therapy. Triggering of NO production, and inhibition of infectivity rate are revealed as mechanisms of action ZnNPs on L. major. But, supplementary investigations are necessary to clear the efficacy and safety of these agents.

Leishmanicidal and immunomodulatory activities of the formononetin (a natural isoflavone) against Leishmania tropica.

OBJECTIVE: This work aimed to examine the leishmanicidal, cellular mechanisms and cytotoxicity effects of formononetin (FMN), a natural isoflavone, against Leishmania tropica. We used the MTT assay to determine the leishmanicidal effects of FMN against promastigotes and its cytotoxicity effects on J774-A1 macrophage cells. The Griess reaction assay and quantitative real-time PCR were used to determine the nitric oxide (NO) and the mRNA expression levels of IFN-γ and iNOS in infected J774-A1 macrophage cells. RESULTS: FMN significantly (P < 0.001) decreased the viability and number of promastigotes and amastigotes forms. The 50% inhibitory concentrations value for FMN and glucantime was 9.3 and 14.3 µM for promastigote and amastigote, respectively. We found that the macrophages exposed with FMN especially at concentrations of 1/2 IC50 and IC50 significantly activated the NO release and the mRNA expression levels of IFN-γ, iNOS. The findings of the current research showed the favorable antileishmanial effects formononetin, a natural isoflavone, against various stages of L. tropica through inhibition of infectivity rate of macrophage cells and triggering the NO production and cellular immunity. However, supplementary works are essential to evaluate the ability and safety of FMN in animal model before use in the clinical phase.

Green Synthesis, Characterization, and Antiparasitic Effects of Gold Nanoparticles against Echinococcus granulosus Protoscoleces.

Echinococcosis, or hydatidosis, is one of the most important zoonotic diseases, which is initiated by the larval stage in the clasts of Echinococcus granulosus. For the treatment of hydatidosis, surgery is still the preferred method and the first line of treatment for symptomatic patients. Unfortunately, most of the scolicidal agents that are injected inside cysts during hydatid cyst surgery have side effects, including leaking out of the cyst and adverse effects on the living tissue of the host, such as necrosis of liver cells, which limits their use. This work was carried out to study the lethal effect of green synthesized gold nanoparticles (Au-NCs) against hydatid cyst protoscoleces. Au-NCs were green synthesized using the Saturja khuzestanica extract. Au-NCs were characterized by UV-visible absorbance assay, electron microscopy analysis, X-ray diffraction (XRD), and Fourier transform infrared (FTIR) spectroscopy. Scolicidal properties of Au-NCs (1-5 mg/mL) were studied against protoscoleces for 10-60 min. The effect of Au-NCs on the expression level of the caspase-3 gene as well as the ultrastructural examination was studied by real-time PCR and scanning electron microscopy (SEM). The cytotoxicity of Au-NCs on hepatocellular carcinoma (HepG2) and normal embryonic kidney (HEK293) cell lines was also studied by the cell viability assay. The obtained Au-NCs are cubes and have an average size of 20-30 nm. The highest scolicidal efficacy was observed at 5 mg/mL with 100% mortality after 20 min of treatment for hydatid cyst protoscoleces. In ex vivo, Au-NCs required more incubation time, indicating more protoscolicidal effects. Au-NCs markedly upregulated the gene level of caspase-3 in protoscoleces; whereas they changed the ultra-structure of protoscoleces by weakening and disintegrating the cell wall, wrinkles, and protrusions due to the formation of blebs. We showed the effective in vitro and ex vivo scolicidal effects of Au-NCs against hydatid cyst protoscoleces by provoking the apoptosis process of caspase-3 activation and changing the ultrastructure of protoscoleces with no significant cytotoxicity against human normal cells. However, additional studies should be conducted to determine the possible harmful side effects and accurate efficacy.

The role of Curcuma longa essential oil in controlling acute toxoplasmosis by improving the immune system and reducing inflammation and oxidative stress.

Background: Chemotherapy with synthetic drugs is the principal approach for toxoplasmosis treatment; however, recent studies reported the limitations and adverse side effects of these chemical drugs. Objective: This study aimed to examine the in vitro and in vivo effects of Curcuma longa essential oil (CLE) against the Toxoplasma gondii RH strain. Methods: The in vitro effect of different concentrations of CLE on T. gondii tachyzoites was assessed by cell viability assay. Flow cytometry and apoptosis analysis were performed, and nitric oxide production by CLE was also evaluated in tachyzoites. BALB/c mice were orally treated with various doses (1.25, 2.5, and 5 mg·kg-1·day-1) of CLE for 2 weeks. After the induction of acute toxoplasmosis in the mice, their survival rate and the mean number of peritoneal parasites were checked. The hepatic level of antioxidant enzymes and oxidative stress markers was evaluated by commercial kits. The mRNA expression level of proinflammatory cytokines such as interleukin 1-beta (IL-1ß) and interferon-gamma (IFN-γ) was evaluated by quantitative real-time PCR. Results: CLE, especially at 50 µg/ml, showed potent inhibitory effects on T. gondii tachyzoites. It increased the survival rate (ninth day) and reduced the mean number of peritoneal tachyzoites in the infected mice. CLE dependently increased (p < 0.01) the number of necrotic and apoptotic cells as well as NO production. CLE significantly (p < 0.05) reduced the hepatic level of oxidative stress markers but increased (p < 0.001) the antioxidant enzymes and proinflammatory cytokines in the infected mice, with no important toxicity for vital organs. Conclusion: The findings of this survey revealed the significant in vitro inhibitory effects of CLE on T. gondii tachyzoites. The results also exhibited promising in vivo effects of CLE. CLE improved the survival rate of infected mice and reduced the parasite number in them. Although the mechanisms of action of CLE are not clear, our study demonstrated its beneficial effects on acute toxoplasmosis by strengthening the immune system and reducing inflammation and oxidative stress. Still, more studies are required to confirm these results.

Molecular epidemiology and associated risk factors of oral cavity parasites in hemodialysis patients in western Iran.

Renal disorders are able to make patients more susceptible to infections, including those caused oral cavity infections. Today, it has been proven that during end stage renal failure, hemodialysis results in acquired immune response defects through failings of humoral and cell-mediated immune. This survey was intended to assess the molecular epidemiology and associated risk factors of Entamoeba gingivalis and Trichomonas tenax as the main oral cavity protozoa in hemodialysis patients in Lorestan Province, western Iran. The investigation was performed on 73 hemodialysis patients referring to hemodialysis centers of Lorestan Province, Iran during May 2021 to February 2022. The frequency of oral cavity protozoa was investigated using microscopic and conventional polymerase chain reaction (PCR). A provided questionnaire with a number of demographical information and related risk factors was completed for each patient. The results showed that E. gingivalis and T. tenax parasites were found in 18 (24.6%) and 20 (27.4%) of the hemodialysis participants by microscopic and PCR test, respectively. Among samples, 13 (17.8%) of the hemodialysis participants were infected with E. gingivalis; whereas 7 (9.6%) of the participants were positive for T. tenax. No significant association was reported between gender, age, education, flossing, use of mouthwash, and prevalence of oral protozoa in hemodialysis participants. However, a significant correlation was observed among between living in rural regions (OR = 13.16; 95% CI = 2.64-56.81; p = 0.002), brushing teeth (OR = 8.51; 95% CI = 1.71-42.3; p = 0.009) and prevalence of oral protozoa in hemodialysis participants. The findings of these epidemiological study clearly showed the high frequency of oral cavity parasites in hemodialysis patients in Lorestan Province, Western Iran. Awareness of the main risk factors for oral cavity parasites particularly teeth brushing is necessary in refining public and oral health policies in hemodialysis patients. Consequently, dental practitioners, nephrologist, and urologist must be alert of these risk factors to carefully identify and achieve oral health concerns in hemodialysis patients to prevent the oral diseases and infections.

High potency of magnetic iron oxide nanoparticles covered by piroctone olamine against cystic echinococcosis.

This study examined the synthesis of magnetic iron oxide nanoparticles coated with PO (FOMNPsP) and assessed their in vitro, ex vivo, and in vivo effects against cystic echinococcosis. The FOMNPsP was synthesized through the alkalization of iron ions in a deoxygenated form. In vitro and ex vivo protoscolicidal effects of FOMNPsP (100-400 µg/mL) were evaluated on hydatid cyst protoscoleces by the eosin exclusion test for 10-60 min. The effect of FOMNPsP on caspase-3 gene expression and exterior ultra-structural of protoscoleces was assessed by real-time PCR and scanning electron microscopy (SEM), respectively. In vivo effects were assessed by evaluating the number, size, and weight of hydatid cysts among infected mice. The FOMNPsP size was < 55 nm, and the most frequent particles were in the 15-20 nm range. In vitro and ex vivo assays revealed that the highest protoscolicidal effect was observed at 400 µg/mL with 100% lethality. After exposure of protoscoleces with FOMNPsP, the level of gene expression of caspase-3 was dose-dependently increased (p < 0.05). By SEM, the FOMNPsP-treated protoscoleces showed wrinkles and bulges resulting from the formation of blebs. FOMNPsP significantly decreased (p < 0.01) the mean number, size, and weight of the hydatid cyst. FOMNPsP revealed the potent protoscolicidal traits through disrupting the cell wall and apoptosis induction. The results also indicated the promising effect of FOMNPsP in controlling hydatid cysts in the animal model. Although FOMNPsP is safe for human normal cells, more investigations are required to clarify its toxicity and precise mechanisms of action.

Iranian Hydatid Disease Registry: Establishment and Implementation of a Neglected Tropical Disease Registry.

BACKGROUND: Cystic echinococcosis (CE) or hydatid disease is a global public health concern which imposes considerable economic costs on the communities in endemic regions. CE surveillance data are not adequately reliable. The present study reports the development and outcomes of a CE registry in Iran. METHODS: Hydatid Registry (HydatidReg) was initially established as a single-center registry in 2014 after the ethical approval of KMU. Following a call from MoHME to promote registry of different diseases and health outcomes, a call for participation was announced and all the Iranian Universities of Medical Sciences were requested to contribute to the registry. Subsequently, a nation-wide registry of hydatid disease was established in 2016. With a global perspective, HydatidReg joined the European Register of Cystic Echinococcosis (ERCE). A data collection form based on minimum dataset was designed and standard operating procedures (SOPs) were prepared to ensure standardized patient enrolment in the registry. A biobank system with two-dimensional barcoding was established along with HydatidReg for management and organization of biological specimens. RESULTS: As of March 2021, a total of 690 patients were enrolled in the registry. HydatidReg registered 362 (17.3%) out of the total 2097 patients enrolled in ERCE. Quality control (QC) of the data demonstrated 91.2% completeness and 80% timeliness. In the biobank, 322 biological specimens from 184 CE patients have been deposited including 70 blood, 96 sera and 156 parasite materials. CONCLUSION: High-quality data in the HydatidReg registry provided opportunities for health professionals to improve quality of care and organize meaningful research.

Anti-helminthic effects and cellular mechanisms of Astragalus ecbatanus extract against Echinococcus granulosus protoscoleces.

We aimed to investigate the in vitro and ex vivo anti-helminthic effects of Astragalus ecbatanus chloroform extract (AECE) as well as its cellular mechanisms against Echinococcus granulosus protoscoleces. In vitro and ex vivo anti-helminthic effects of AECE on E. granulosus protoscoleces were evaluated through eosin exclusion test. Effects of AECE on induction of the caspase-3 like activity, the permeability of plasma membrane were also determined. The effects of AECE against the human hepatocyte carcinoma (HepG2) and human embryonic kidney (HEK293T) cells were determined by MTT ([3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyl tetrazolium bromide)]) test. The findings revealed that AECE especially at 45 mg/mL displayed potent in vitro and ex vivo anti-helminthic effects against E. granulosus protoscoleces (p < 0.001). After treatment of protoscoleces with AECE, the caspase-3 enzyme activity and the rate of permeability of plasma membrane was dose dependently (P < 0.001) increased. The 50% cytotoxic concentration (CC50) value of AECE for normal and cancer cells lines 4.62 mg/mL and 1.89 mg/mL, respectively. The findings of the current study revealed the favorable in vitro and ex vivo anti-helminthic effects of AECE against protoscoleces of E. granulosus. However, further surveys in animal model are required to elucidate different aspects of this extract before use in clinical phases.

Potential effects of alpha-pinene, a monoterpene commonly found in essential oils against Toxoplasma gondii infection; an in vitro and in vivo study.

This survey designed to assess the in vitro and in vivo activity of α-pinene, a monoterpene commonly originated in essential oils on Toxoplasma gondii. The in vitro effect of various concentration of α-pinene against tachyzoites of T. gondii Rh strain was assessed by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. The activity of α-pinene on the stimulation of apoptosis in tachyzoites of T. gondii was also examined using the caspase 3 colorimetric activity assay. In vivo assay, mice were orally received α-pinene at 2 and 4 mg/kg/day for 14 days, then, pre-treated mice were daily tested and the rate of death was recorded. α-pinene meaningfully declined (p < 0.001) the tachyzoites viability with the IC50 value of 23.3 µg/mL. α-pinene induced the apoptosis through increasing the caspase-3 activity by 35.6%. Oral treatment with α-pinene significantly (p < 0.01) improved the survival rate infected mice with by 9th day. α-pinene + atovauone (50 mg/kg) significantly (p < 0.01) improved the survival rate infected mice up to 11 days compared with the control groups. α-pinene especially in combined atovaquone at 50 mg/kg for 2 weeks meaningfully (p < 0.05) declined oxidative stress. We found the promising in vitro anti-Toxoplasma effects of α-pinene on T. gondii RH strain. In addition, we found that α-pinene therapy particularly along with the reference drug declined the mortality rate of infected mice. Although, we just confirmed the stimulation of apoptosis and anti-inflammatory effects as the main anti-Toxoplasma mechanisms of α-pinene; however, more surveys concerning the accurate mechanisms, toxicity, and efficacy on other T. gondii strains are required to confirm these results.

Prevalence and risk factors of oral cavity parasites in pregnant women in Western Iran.

Background: We aimed to study the frequency and risk factors of oral cavity protozoa (Trichomonas tenax and Entamoeba gingivalis) in pregnant women in Lorestan province, western Iran. Methods: The current cross-sectional descriptive work was performed on 200 pregnant women referring to health centers of Lorestan Province, Iran during August 2021 to February 2022. Two specimens were obtained from each women by means of sterile swabs from saliva and dental plaques. The frequency of oral cavity protozoa was investigated using microscopic and conventional polymerase chain reaction (PCR). Results: E. gingivalis and T. tenax parasites were found in 41 (20.5%) and 46 (23%) of the pregnant women by microscopic and PCR test, respectively. Among positive samples, 29 (63.1%) of the pregnant women were infected with E. gingivalis; whereas 19 (36.9%) of the participants were positive for T. tenax. In the multivariate model, living in rural regions (p < 0.001) and brushing teeth (p = 0.021) were considerably linked with the prevalence of oral cavity parasites. Conclusion: The current study evidently exhibited the high frequency of oral cavity parasites (E. gingivalis and T. tenax) in pregnant women in Lorestan province, Western Iran. Awareness of the main risk factors for oral cavity parasites especially teeth brushing is necessary in refining public and oral health approaches in pregnant women. Thus, dental practitioners and gynecologists must be alert of these risk factors to carefully identify and manage oral health concerns in pregnant women to prevent the oral diseases and infections.

Chemical composition, antileishmanial, and cytotoxic effects Ferula macrecolea essential oil against Leishmania tropica.

Background: The current study was aimed to evaluate the chemical composition, as well as antileishmanial and cytotoxic effects of the essential oil of Ferula macrecolea and its main compound, terpinolene, against promastigotes and amastigotes of Leishmania tropica. Methods: The chemical composition of the essential oil was analyzed by a gas chromatograph connected to a mass spectrometer (GC/MS). The MTT (3-(4.5-dimethylthiazol-2-yl) - 2.5-diphenyl tetrazolium bromide) assay was used to study the effects of the essential oil and terpinolene against promastigotes while the macrophage model was used for evaluating the effect of F. macrecolea essential oil against amastigotes of L. tropica as well as assessing cytotoxicity. The Griess reaction assay was employed to study the nitric oxide (NO) produced by treating macrophage cells with the essential oil and terpinolene. Furthermore, the effect of the essential oil and terpinolene on plasma membrane permeability and inhibition of infection in macrophages was evaluated. Results: The main compounds were terpinolene (77.72%), n-nonanal (4.47%), and linalool (4.35%), respectively. The 50% inhibitory concentrations (IC50) of the essential oil, terpinolene, and glucantime against promastigotes were 27.6, 11.6, and 32.8 µg/mL, respectively; however, their IC50 values against amastigotes were 42.3, 19.6, and 56.9 µg/mL, respectively. The 50% cytotoxic concentrations of the essential oil, terpinolene, and glucantime were 471.3, 207.3, and 1165.3 µg/mL, respectively. The production of NO in macrophage cells after treatment with the essential oil and terpinolene was increased in a dose-dependent manner (p < 0.001). The results revealed that by increasing the concentration of the essential oil and terpinolene, the permeability of the parasites' plasma membrane was significantly changed (p < 0.001). The pre-incubation of Leishmania parasites with F. macrecolea essential oil and terpinolene significantly declined the rate of cell infection by 74.8% and 79.4%, respectively (p < 0.001). Conclusion: The results of the present study indicated that F. macrecolea essential oil, especially its main compound, i.e., terpinolene, has a potent antiparasitic effect on the promastigote and amastigote stages of L. tropica. Considering the advantages of medicinal plant products over their chemical counterparts, it is suggested that in the continuation of this study, the effect of F. macrecolea essential oil, especially terpinolene, on laboratory animals, and in case of high efficiency, in humans be evaluated.

High prevalence of Cryptosporidium infection in Iranian patients suffering from colorectal cancer.

The present investigation was designed to study the prevalence of cryptosporidiosis in the colorectal cancer patients compared to the healthy subjects. The present descriptive case-control study was performed on 174 subjects including 87 healthy people and 87 patients with colorectal cancer attending to general hospitals in Lorestan Province, Western Iran, during October 2019-August 2020. A fresh stool specimen was collected from each subject in a sterile labeled container. The collected stool samples were concentrated using the sucrose flotation method and then prepared for Ziehl-Neelsen staining for microscopic examination. All samples were also tested using the Nested-PCR assays by amplifying the 18S rRNA gene for the presence of Cryptosporidium DNA. Demographic and possible risk factors such as age, gender, residence, agriculture activity, history of contact with livestock, consumption unwashed fruits/vegetables, and hand washing before eating were investigated in all the studied subjects using a questionnaire. Of the 87 patients with colorectal cancer, 37 (42.5%) had Cryptosporidium infection. A significant difference (p < 0.001) in the prevalence of Cryptosporidium spp. infections among the participants in the case and control (11, 12.6%) groups was observed. We found that cryptosporidiosis was not linked with age, gender, hand washing, agriculture activity, and history of contact with livestock in the colorectal patients. However, residence in urban areas was significantly associated with the prevalence of cryptosporidiosis. The 18 s rRNA gene of Cryptosporidium in 48 samples was successfully amplified by the Nested-PCR. Based on the obtained findings, Cryptosporidium spp. infections were observed significantly more frequently in the patients with colorectal cancer in comparison with the healthy individuals. It is suggested to carry out similar studies in various parts of Iran with larger sample sizes and further parasitological tests.

Effects of green synthesized zinc nanoparticles alone and along with albendazole against hydatid cyst protoscoleces.

Background: The current investigation aims to green synthesized the zinc nanoparticles (ZnNPs) using Lavandula angustifolia extract by microwave technique and its protoscolicidal effects alone and combined with albendazole against hydatid cyst protoscoleces. Methods: Different concentrations of the ZnNPs (50, 100, and 200 µg/ml) alone and combined with albendazole (ALZ, 100 µg/ml) were treated with hydatid cyst protoscoleces obtained from liver of infected sheep for 5-60 min in vitro and ex vivo. Eosin exclusion examination was used to assess the viability of protoscoleces. The induction of apoptosis in hydatid cyst protoscoleces was assessed by measurement of the Caspase-3 activity of protoscoleces treated with various concentrations of ZnNPs. Results: The size of green synthesized ZnNPs was ranged from 30 to 80 nm, most of these nanoparticles were between 50 and 60 nm in size. In vitro, the highest scolicidal effect of ZnNPs was observed at the concentration of 200 µg/ml, where it killed 81.6% of protoscolices. While the combination of these nanoparticles with ALZ, especially at the concentration of 200 µg/ml, completely killed the protoscolices after 10 min' exposure. However, compared to in vitro assay, the drugs tested took longer to show their protoscolicidal effect. Conclusion: Based on the obtained results, ZnNPs particularly in combination with albendazole displayed the potent protoscolicidal in vitro and ex vivo as an intraperitoneal model of administration of agents to hydatid cyst treatment; nevertheless, additional investigations are mandatory to evaluate the efficacy and safety Zn NPs as a favorable protoscolicidal agent in clinical setting.