In this review, we intend to provide a summary of the activities of researchers in the field of Toxoplasma gondii in Iran, during the past 70 years. Most studies have been limited to epidemiological studies (mostly using ELISA and IFA methods). Designing a standard and reliable method using the specific antigens of this parasite is essential. So far, studies in the field of drug effects have not been able to introduce an effective drug with few side effects. Various types of vaccines have been developed, such as recombinant and DNA vaccines. However, none of them had a good efficacy. The use of multi-epitope vaccines as potential vaccines against toxoplasmosis is recommended. At present, limited studies have been conducted on the patterns of transmission and genetic diversity of isolated isolates in Iran. Future research to determine the genotype of T. gondii could play an important role in the study of population structure, and biological characteristics of this parasite. It is hoped that the results of this study will help control, prevent, and reduce the burden of disease caused by this parasite.

Acanthamoeba is a free-living amoeba that has been found on surfaces, air, water and various environmental sources around the world. It enters the human body through the respiratory tract via the nose. Of note, amoebae are well known to act as a reservoir for various pathogenic microorganisms including bacteria and viruses such as Adenoviruses and Mimivirus. Given that SARS-CoV-2 of the Coronaviridae family is transmitted through the respiratory tract, and the Trojan horse nature of Acanthamoeba, it has been suggested that amoebae act as a vector in the transmission of SARS-CoV-2. The aim of this study was to provide an opinion of the possibility of the coexistence of Acanthamoeba with SARS-CoV-2.

Background: Visceral leishmaniasis (VL) is a lethal parasitic disease, transmitted by sand fly vectors. Immunomodulatory properties of sand fly saliva proteins and their protective effects against Leishmania infection in pre-exposed animals suggest that a combination of an antigenic salivary protein along with a Leishmania antigen can be considered for designing a vaccine against leishmaniasis
Methods: Three different fusion forms of L. infantum hypothetical protein (LiHyV) in combination with Phlebotomus kandelakii salivary apyrase (PkanAp) were subjected to in-silico analyses. Major Histocompatibility Complex (MHC) class I and II epitopes in both humans and BALB/c mice were predicted. Antigenicity, immunogenicity, epitope conservancy, toxicity, and population coverage were also evaluated. Results: Highly antigenic promiscuous epitopes consisting of truncated LiHyV (10-285) and full-length PkanAp (21-329) were identified in human and was named Model 1. This model contained 25 MHC-I and 141 MHC-II antigenic peptides which among them, MPANSDIRI and AQSLFDFSGLALDSN were fully conserved. LALDSNATV, RCSSALVSI, ALVSINVPL, SAVESGALF of MHC-I epitopes, and 28 MHC-II binding epitopes showed 60% conservancy among various clades. A population coverage with a rate of >75% in the Iranian population and >70% in the whole world was also identified.
Conclusion: Based on this in-silico approach, the predicted Model 1 could potentially be used as a vaccine candidate against VL.

Background: Fleas (Insecta: Siphonaptera) are considered as highly specialized bloodsucking on mammals such as humans, livestock, dog, cat, rabbit, squirrels, rats, and mice. The desire for blood feeding from warm-blooded animals has led to becoming an intermediate host for some tapeworms like Dipylidium. The aim of this study was to detect D. caninum larval infection in fleas of dogs living in Meshkinshahr County, northwest of Iran.
Methods: Fleas were collected from 42 dogs using brushing the hair in Meshkinshahr for one year (2014-2015). After the morphological study, fleas were preserved in 90% ethanol for molecular identification. After DNA extraction, the 28S ribosomal RNA gene (~670 bp) of D. caninum was amplified using specific primers. Finally, the PCR products were sequenced.
Results: Overs, 974 fleas were collected from the dogs. In the morphological study, three species Ctenocephalides canis, Ct. felis, and Pulex irritans were identified. PCR and sequence analysis results showed that 4 isolates Ct. Canis were infected with D. caninum. Also, no positive specimens were isolated from the other two species.
Conclusion: Ct. canis is reported as the most important species of fleas in transferring D. caninum in that region.

Background: Free-living amoebae (FLA) such as Acanthamoeba spp., are considered as opportunistic and pathogenic protozoans.  Acanthamoeba granulomatous encephalitis (AGE) is a serious threat for immunodeficient patients and Acanthamoeba keratitis (AK) for contact lens users. We aimed to identify the presence of free living amoebae in nasal swabs of patients and contact lens users in Qazvin, Iran.
Methods: During 2019, 251 nasal and oral swabs (including the pharynx and mouth) were collected from patients with diabetes, AIDS and those under periodic dialysis in Qazvin, Iran. In addition, 27 soft contact lenses were collected from the participants. Following DNA extraction, PCR and sequencing were conducted to identify the genotypes of the amoeba. Phylogenetic analysis of the identified sequences was performed using MEGA 7 software.
Results: A strain of Acanthamoeba belonging to the T3 genotype was isolated from hemodialysis patients. Two specimens of Acanthamoeba with T3 genotype were isolated from keratitis patients.
Conclusion: The clinicians should pay attention to the possible complication of this organism because this amoeba is potentially pathogenic for immunocompromised patients. Since the amoeba is present in environmental resources, the use of contact lenses should be accompanied by considering proper hygiene.

Background: The risk of transmission of some infectious agents has always been an important life-threatening side effect of blood transfusion. The aim of this study was to develop a triplex nested PCR (tnPCR) to assess the presence of protozoan parasites Plasmodium, Toxoplasma and Babesia in blood samples concurrently. The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of developed tnPCR method compared to gold standard methods were determined for all three parasite genera.
Methods: After selecting the genus level specific primers and setting up of tnPCR, blood samples were collected partially from Isfahan Blood Transfusion Organization (IBTO). Some different samples from human and animal were tested by this method in comparison with the gold standard methods (microscopic method for Plasmodium, Babesia and ELISA for Toxoplasma) in 2021.

Results: This tnPCR works well and the sensitivity, specificity, PPV, NPV and accuracy, of this molecular method were all 100% for Plasmodium spp., 93.33%, 99.16%, 93.33%, 99.16% and 99.25% for Babesia spp. and 100%,98.5%, 85.72%, 98.5% and 100% for Toxoplasma gondii respectively compared to standard methods. In average there were 100%, 99.22%, 95.24%, 99.5% and 99.75% contingency for all three parasites (α<0.05). The designed and provided method can detect one, two, or all three potentially dangerous pathogens simultaneously in one tube and one-step, in biological specimens as well as blood.
Conclusion: The developed tnPCR worked well. It could be recommended for facilitating test, saving time, reducing the expense and cross contamination, subsequently the promotion of blood transfusion safety.

Background: Blastocystis sp., is a prevalent protist isolated from humans and animals, which its opportunistic role in immunocompromised patients is still controversial. The current study aimed to evaluate the subtype and alleles distribution of Blastocystis sp., among immunocompromised patients.
Methods: Totally, 33 microscopically Blastocystis-positive stool samples, isolated from Guilan province during April 2018 to May 2019 were investigated. Total DNA extraction was performed and the barcoding region of the small subunit ribosomal RNA (SSU rRNA) gene was amplified. Targeted fragments were sequenced to characterize subtypes and relevant alleles. Phylogenetic tree was constructed using Maximum-likelihood and Tamura 3-parameter to illustrate the correlation between subtypes and certain immunodeficiency.

Results: Subtype analysis revealed the presence of ST1, ST2, ST3, and ST7 among 13/33 (39.4%), 5 (15.2%), 14/33 (42.4%), and 1/33 (3%), of samples, respectively. ST1 was the major subtype among cancer patients 5/7 (71.42%), while ST3 was the predominant subtype among rheumatoid arthritis (RA) patients 3/6 (50%), internal ward patients 5/10 (50%), and asthma and allergy patients 2/3 (66.66%). ST7 was isolated from a patient hospitalized in internal ward. No significant correlation was seen between the type of immunodeficiency and subtypes (P-value = 0.771). The phylogenetic tree showed no separation regarding the type of immunodeficiency.
Conclusion: Among studied immunocompromised patients, ST3 was the most prevalent subtype followed by ST1. There was no specific correlation between subtypes and alleles with type of immunodeficiency. Putative zoonotic alleles were highlighted the probability of zoonotic transmission for Blastocystis sp.

Background: Two calcified objects recovered from an adolescent in a burial site in Amiens, France, have been previously identified as hydatid cysts using thin-section petrography. The importance of ancient hydatidosis besides the value of these unique archeological excavated materials encouraged the authors to look at this attractive subject more interdisciplinary by implementing medical radiology.
Methods: In the current experiment, which has been carried out in the Radiology Department, Tehran Heart Center (THC), Tehran, Iran, the conventional and dual-energy dual-source CT-scan was used in studying the remaining structures of the two calcified masses. The imaging procedure was carried out based on X-Ray attenuation by two different tube voltages.
Results: A high concentration of calcium sediment in the cyst walls was revealed in Hounsfield units, the measuring of the elements in CT. Taking advantage of implementing this imaging technique the oxalate calcium was also shown as the dominant component of the samples. The results were all in favor of diagnosing hydatid cysts.
Conclusion: The achieved pictorial results in the present paper have highlighted the important role of CT scan as a noninvasive confirming technique in paleopathological investigations. Using Dual-source dual-energy CT-scan in reconfirming these previously identified hydatid cysts, is an encouraging message towards the necessity of sequential studies on invaluable biological excavated pieces.

Background: The application of plumbagin (PLN), with a wide use in pharmaceutical science, is limited due to its low water solubility and poor bioavailability. Micelles can encapsulate hydrophobic drugs due to their hydrophobic core. The aim of this study was to develop and characterize a polymeric micelle formulation of PLN and evaluate its in vivo anti-plasmodial property.
Methods: The study was conducted at Zanjan University of Medical Sciences, Zanjan, Iran in 2018. The triblock copolymeric micelles of PLN was prepared by e-caprolactone ring-opening polymerization, by PEG as the macroinitiator and using Sn(Oct)₂ for its catalytic properties . The synthesized nanoparticles were characterized by ¹H NMR, FTIR, GPC, AFM, and DLS. The encapsulation efficiency, drug loading capacity, and drug release were measured by UV-Vis at 520 nm. Also in vivo anti-plasmodial potential of fabricated drug loaded micelle was investigated using the 4-day suppressive test against Plasmodium berghei infection in mice.
Results: The nanoparticles average diameter was obtained less than 80 nm. The loading capacity and encapsulation efficiencies were 18.9±1.3% and 81±0.78%, respectively. In vitro, PLN release studies showed a sustained-release pattern until 7 days in PLN-loaded micelles (M-PLN) and drug release rate in acidic condition was higher than neutral condition. In vivo, anti-plasmodial results against P. berghei displayed an 8-fold increase in anti-plasmodial activity of M-PLN when compared to free PLN at the tested dosage level on the 7^th day.
Conclusion: Based on these results, PCL–PEG–PCL micelles have a great potential to be the carrier for PLN for the malaria targeting.

Background: This study was conducted to determine the prevalence and intensity of nematode infections in sheep located in northeastern Iran from Apr 2018 to Mar 2019.
Methods: Gastrointestinal nematodes of 300 sheep were inspected. The season of slaughter, anatomic location where the parasite was located, the animal’s sex, infection prevalence and intensity were recorded. Seasonal differences in arrested larvae numbers also were assessed using Cochran's Q test.
Results: Overall, 4,331 adult nematode specimens were collected. Among the examined sheep, 53% (159/300) were infected with one or more nematode species. Among infected sheep, 42.8% were infected with a single species of nematode, 26.4% were infected with two species of nematodes, and 30.8% were infected with three or more species of nematodes. Marshallagia marshalli (13.3%) was the most common nematode recovered from the abomasums of infected sheep, while Trichostrongylus vitrinus (4.6%) was commonly recovered from the small intestines, and Trichuris ovis (25.6%) was commonly recovered from the large intestines. In total, 463 arrested larvae were found in the abomasums of 7.5% of infected sheep and 104 arrested larvae were found in the small intestines of 8.8% of infected sheep. A significantly higher numbers of arrested larvae were found in summer compared to autumn (P<0.001).
Conclusion: Intestinal parasites continue to be a problem for sheep in northeastern Iran and additional control measures need to be explored.

Background: We aimed to evaluate currently protoscolices agents by using two methods including injection and impregnated sponges with protoscolices agents.
Methods: Hypertonic saline 20%, silver nitrate 0.5% and Ethanol 96% were injected into sheep hydatid cysts. Then, after different exposure times (5, 10, and 15 min), the protoscolices were aspirated from the cyst and viability of the protoscolices were evaluated microscopically using vital staining. In the second part, the live protoscolices were sprayed on the soaked sponge with the mentioned protoscolicidal agents. After different exposure times live protoscolices were counted microscopically. One-way ANOVA and Tukey tests were used for showing difference significant between studied groups.
Results: Cyst injection method showed that ethanol, Nacl and AgNo3 have 80%, 60% and 6% protoscolocidal efficacy, respectively (P<0.05). In sponge method the most and fastest protoscolocidal efficacy were observed in all used protoscolicidal agents.
Conclusion: Ethanol and NaCl and AgNo3 with sponge method is more effective than the injection of protoscolices agents in the cyst.

Background: We aimed to estimate the incidence of Toxoplasma infection in T. gondii-seropositive patients under allogeneic hematopoietic stem cell transplantation (HSCT).
Methods: The present research was a prospective study on 54 whole blood samples of allogeneic HSCT recipients, who were referring to bone narrow transplantation centers affiliated to Shahid Beheshti University of Medical Sciences, Tehran, Iran in 2018. All patients were IgG positive against T. gondii.
Results: Overall, 54 Toxoplasma positive pre-HCTSP patients were enrolled. 53.7% (n= 29) were male, also 1.9% (n=1) had germ-line type of the disease. The Multiple myeloma patients had higher age in comparison with other disease, but pairwise comparison showed the difference of age between Multiple myeloma patients were statistically significant with Acute lymphoblastic leukemia, Acute myeloblastic leukemia and Huntington's disease (P< 0.05). The results of PCR assay showed 5.6% (n= 3) of the patients were infected with Toxoplasma.
Conclusion: PCR method has detected considerable incidence of Toxoplasma infection for monitoring HSCT recipients at risk for toxoplasmosis, and many patients who showed the incidence of toxoplasmosis had previous infections with the Toxoplasma parasite.

Background: Studies on experimental model of cancer showed that hydatid cyst fluid (HCF) has antitumor activity. We aimed to investigate the effect of HCF and Antigen B (AgB) on 4T1 breast tumor cells in BALB/c mice.
Methods: This study was carried out in the Department of Parasitology, Tarbiat Modares University, Tehran, Iran from 2019 to 2020. There were two control groups of BALB/c mice (one group were injected with aluminum sulfate and another group with PBS), and six groups, injected via the intraperitoneal route with 100, 300 and 500 μg/ml concentrations of HCF, AgB diluted in 100 μl PBS, and alum. Seven days after the last treatment, 7×10⁵ 4T1 cells were subcutaneously injected into the right flank of BALB/c mice.
Results: The difference between the mean size of the tumor in the case and control groups was statistically significant (P<0.05). The mean level of cytokines between the case and control groups was statistically significant (P<0.05). Our histopathological studies showed a reduction in both tumor volume and carcinogenesis in groups injected with 300 μg/ml HCF and 500 μg/ml AgB. The antitumor activity of HCF and AgB may be related to the immune responses against these antigens.
Conclusion: We suggest that polarization of the Th1/Th2 ratio toward the Th1 pathway occurred in groups injected with HCF and AgB. More comprehensive and precise experiments using different hydatid cyst components are required to investigate their prophylactic effects on breast cancer.

Background: Echinococcosis is a common parasite with zoonotic character created by a small cestode, Echinococcus spp., and is an important public health problem in Turkey as well as all over the world. We aimed to investigate antibodies in serum samples of suspected Echinococcosis patients sent to the National Parasitology Reference Laboratories of the General Directorate of Public Health.
Methods: Serum samples of 2390 patients sent to our laboratory between January 1, 2014 and May 01, 2019, evaluated by ELISA, Indirect Hemagglutination Test (IHA) and Western Blot (WB) methods are presented. Our laboratory is the national reference laboratory. All kinds of tests requested from suspected patients can be performed
Results: Overall, 1199 (50.2%) of 2390 serum samples were female and 1191 (49.8%) were male. It was observed that 178 (14.9%) of men and 210 (17.5%) of women were seropositive. There was no statistical difference between the sexes in terms of seropositivity. Of all samples, 1941 (81.2%) were negative, 388 (16.2%) were positive, and 61 (2.6%) were borderline. Results determined as borderline are considered suspicious and a recommendation is made to repeat the test after 15 days. A statistical difference was found in the distribution of seropositivity by years. While seropositivity was lowest in 2014, it was found to be highest in 2018 and 2019.
Conclusion: Despite all the precautions taken, it is seen that echinococcosis still continues to exist in Turkey as a zoonotic disease. Hence, CE has been involved in Turkey Zoonotic Diseases Action Plan (2019-2023) and decided to carry out studies for the protection and prevention of the disease.

Background: The gastrointestinal parasite Giardia lamblia causes giardiasis. Its treatment with standard drugs produces side effects and improper treatment can generate resistant strains. New antigiardial compounds are needed. An analysis was done to identify the antigiardial activity of Morinda royoc, a plant used in traditional Mayan medicine to treat stomach and bowel pain. We aimed to assess the efficacy of M. royoc roots against G. lamblia and their effect on cells viability.
Methods: A methanol extract was done of the root and then fractionated. The extract and fractions were tested in vitro on G. lamblia trophozoites and their effect on cell viability was quantified by flow cytometry. The active extract and fractions were analyzed by gas chromatography–mass spectrometry and high-performance liquid chromatography.
Results: The hexane fraction exhibited potent activity against G. lamblia (IC₅₀ = 0.08 µg/mL). Its principal component was an anthraquinone-type compound. None of the fractions were toxic to human promyelocytic leukemia, chronic myelogenous leukemia and human mononuclear cells.
Conclusion: The medicinal plant M. royoc contains promising bioactive agents with antigiardial activity and deserves further research.

Background: Freshwater snails serve as intermediate hosts for a variety of trematodes that cause illness in the human and animal populations. Several species of freshwater snails in Thailand have been found to have larval trematode infections. We aimed to investigate a freshwater snail in Phitsanulok Province and report on its current status of larval trematode infection.
Methods: Freshwater snails were collected from six localities (rice field and irrigation canal) by handpicking and using a count per unit of time sampling approach. The snails were identified by their external shell morphology. The shedding method was applied to observe the cercariae, which were photographed under a light microscope to determine their morphological types.
Results: A total of 211 snails were classified into seven genera. The most abundant snail species was Lymnaea sp., representing 31.3% of the sample, followed by Physella sp., Bithynia sp., Pomacea canaliculata, Filopaludina martensi, Indoplanorbis exustus, and Melanoides tuberculata, in that order. From the sample, 21 snails (9.95%), including Bithynia sp., Lymnaea sp., I. exustus, and M. tuberculata, were infected with cercarial trematodes, which could be categorized into four types, namely amphistome, parapleurolophocercous, echinostome, and xiphidiocercaria. Amphistome emerged from Bithynia sp., and I. exustus was the most common cercaria to be recovered, representing 80.9% of all infected snails.
Conclusion: This study presents the current prevalence of cercariae in infected snails within the studied area. It is important to manage intermediate host snails in order to restrict trematode life cycle completion.

Hydatid disease (HD) is a zoonotic disease, which typically affects the liver and the lungs. Primary retroperitoneal localization of the disease is infrequent. The most common symptom is abdominal pain; however, the clinical features of HD may be generally dependent on the location of the cyst. 
A 53-year-old female patient presented to Kocaeli University General Surgery Clinic, Turkey in September 2019 with complaints of flank pain, constipation and vomiting. The MRI revealed cystic lesions in the pancreas and psoas muscle. Both cysts were excised totally during laparotomy. Histopathology reports showed that the lesion in psoas muscle was hydatid cyst whereas the lesion in pancreas was pancreatic pseudocyst. Postoperative period was uneventful and the patient was discharged with albendazole treatment. No recurrence occurred during follow-up period. Atypical localization for hydatid disease is a diagnostic dilemma for surgeons, especially in endemic countries. Hydatid disease should always be considered in the differential diagnosis of retroperitoneal lesions, inadequate management can lead to complications and fatalities. Surgery combined with antiparasitic therapy, may be an option in the presence of obstruction of gastrointestinal and/or urinary tract.

Trichuris vulpis is a relatively common cause of canine large bowel disease leading to chronic diarrhea. Faecal analysis is negative most of the times due to long prepatent period and eggs may be shed intermittently. This case study done at Madras Veterinary College, Chennai, Tamil Nadu, India from 2014 to 2018 describes the usefulness of endoscopy in diagnosing the colitis due to presence of T. vulpis worms. Three dogs presented with haematochezia and tenesmus were subjected to detailed clinical examination. As the faecal examination was negative and there was no improvement on initial therapy endoscopy was performed. Numerous T. vulpis whipworms were found attached to the colon mucosa on endoscopic examination. Multiple bleeding ulcerated sites were observed throughout the colon. The dog was treated with oral febental, praziquantal and pyrantal combination tablets (Drontal Plus). Clinical improvement was observed within a week.