A global systematic review and meta-analysis on the babesiosis in dogs with special reference to Babesia canis.

BACKGROUND: Canine babesiosis is a clinically significant tick-transmitted disease caused by several species of the intraerythrocytic protozoan parasite Babesia, which result in a wide range of clinical manifestations, from mild, transient infection to serious disease and even death. OBJECTIVES: The current study aimed to estimate the global prevalence and associated risk factors of Babesia in dogs. METHODS: Multiple databases (PubMed, Scopus, ProQuest, Web of Science and Google Scholar) were searched for relevant literature published from January 2000 up to December 2022. The statistical analyses were performed based on the R software (version 3.6) meta-package. RESULTS: Out of 23,864 publications, 229 studies met the inclusion criteria. The pooled prevalence of canine babesiosis was 0.120 (95% CI; 0.097-0.146). The highest pooled prevalence was found in Europe (0.207, 95% CI; 0.097-0.344). Among several species, Babesia canis was the most prevalent parasite (0.216, 95% CI; 0.056-0.441). The highest pooled prevalence of Babesia in dogs was observed in the summer season (0.097, 95% CI; 0.040-0.174). CONCLUSIONS: Regular screening and appropriate control strategies are recommended for the prevention of transmission of tick-borne disease transmission among dogs.

In Vitro Evaluation of Anti-Parasitic Activities of Quinolone-Coumarin Hybrids Derived from Fluoroquinolones and Novobiocin Against Toxoplasma gondii.

PURPOSE: Toxoplasmosis is caused by the parasite Toxoplasma gondii (T. gondii). In immunocompetent individuals, the infection is often asymptomatic; however, in expectant mothers and those with immune system deficiencies, complications may arise. Consequently, there is a need for new drugs that cause minimal damage to host cells. The purpose of this study was to investigate the in vitro antiparasitic efficacy of quinolone-coumarin hybrids QC1-QC12, derived from quinolone antibacterials and novobiocin, against T. gondii. METHODS: The derivatives were compared with novobiocin and ciprofloxacin during testing, with pyrimethamine used as a positive control. We conducted the MTT assay to examine the anti-toxoplasmic effects of the test compounds and novobiocin. Evaluation included the infection and proliferation indices, as well as the size and number of plaques, based on the viability of both healthy and infected cells. RESULTS: The in vitro assays revealed that QC1, QC3, QC6, and novobiocin, with selectivity indices (SIs) of 7.27, 13.43, and 8.23, respectively, had the least toxic effect on healthy cells and the highest effect on infected cells compared to pyrimethamine (SI = 3.05). Compared to pyrimethamine, QC1, QC3, QC6, and novobiocin Without having a significant effect on cell viability, demonstrated a significant effect on reducing in both infection index and proliferation index, in addition to reducing the quantity and dimensions of plaques ( P < 0.05). CONCLUSION: Based on our results, QC1, QC3, QC6, and novobiocin due to their significant therapeutic effects could be considered as potential new leads in the development of novel anti-Toxoplasma agents.

First molecular insights into gastrointestinal helminths of domestic birds in the Caspian Sea Littoral of Iran with an emphasis on the One Health concern.

The current investigation was carried out during the period from July 2022 to March 2023, aiming to investigate the prevalence of gastrointestinal helminths in domestic birds collected from traditional markets in Guilan province. One hundred forty-eight domestic birds, including chickens (Gallus gallus domesticus), domestic ducks (Anas platyrhynchos domesticus), greylag geese (Anser anser), and domestic turkeys (Meleagris gallopavo domesticus) were examined. Totally, 42.56% of the investigated birds were positive for helminthic parasites. Morphological analysis revealed varying infection rates among birds: Echinostoma revolutum (5.40%), Hypoderaeum conoideum (2.02%), Cloacotaenia megalops (0.67%), Hymenolepididae family (4.05%), Ascaridia galli (16.89%), and Heterakis gallinarum (4.72%). The investigation involved molecular analysis of the 18S and ITS1 + 5.8S + ITS2 rRNA gene regions. The findings indicated that the 18S region of nematode isolates exhibited a similarity of 92 to 100% with sequences in the GenBank, whereas trematode and cestode isolates showed a gene similarity ranging from 88 to 99%. The ITS regions of nematode, trematode, and cestode isolates exhibited genetic similarities ranging from 87 to 100%, 73-99%, and 75-99%, respectively. Furthermore, phylogenetic analysis confirmed the categorization of the identified species within the Ascaridiidae, Heterakidae, Hymenolepididae, and Echinostomatidae families, indicating their close affinity with previously documented species. Implementing precise control measures such as consistent monitoring, adequate sanitation protocols, and administering anthelmintic treatments is crucial for effectively managing parasitic infections in free-range and backyard poultry farms. Additionally, conducting further surveys is advisable to assess the impact of these parasites on the health and productivity of poultry in the investigated area.

Exploring the Relationship Between KRAS, NRAS, and BRAF Mutations and Clinical Characteristics in Iranian Colorectal Cancer Patients.

BACKGROUND: Patients with colorectal cancer can benefit from anti-EGFR (epidermal growth factor receptor) therapy. However, this therapy is not effective for treating colorectal cancers with constitutive activating mutations in the KRAS, NRAS, and BRAF genes. Molecular analysis of tumor tissue frequently informs treatment decisions for colorectal cancer. This study aims to identify KRAS, NRAS, and BRAF mutations in Iranian patients diagnosed with colorectal cancer and to assess the prevalence of these mutations relative to the tumor differentiation stage within these populations. METHODS: From April 2018 to December 2022, 2000 specimens from patients with colorectal cancer were collected. Data on sex, age, and tumor differentiation stage were recorded for all samples. For mutation detection, the KRAS and NRAS exons (2, 3, and 4) were amplified using the Diatech kit, and a specific primer was used to amplify BRAF exon 15. Pyrosequencing was then performed. RESULTS: Analysis of samples revealed that 1105 specimens (55.3%) contained mutations in at least one of the screened genes. Among the genes studied, the highest occurrence was the KRAS mutation at 47.4%, followed by NRAS at 5.3% and BRAF at 2.7%. Most KRAS mutations were found in exon 2 (89.7%), with the G12D mutation being the most prevalent at 32% of cases. There was a significant difference in the rate of KRAS mutations in women (52.5%) compared to men (43.5%) (P = 0.02). For NRAS, the majority mutations were observed in exon 3 (76.2%), with the Q61H mutation being the most prevalent at 28.5% of cases. There were no significant associations between the clinicopathological parameters and mutations. CONCLUSION: The study's findings indicate a rising frequency of mutations in these genes in Iran, highlighting the need to screening mutations in the main exons of all three genes for effective colorectal cancer treatment strategies.

Biochemical Properties and Immunogenic Epitopes of Echinococcus granulosus Glutathione S-Transferase as a Vaccine Target: In-Silico Study.

Background: The current in silico study was done to determine the primary biochemical features and immunogenic epitopes of Echinococcus granulosus glutathione S-transferase protein as a potential vaccine candidate. Methods: Several web tools were employed to predict physico-chemical properties, antigenicity, allergenicity, solubility, post-translational modification (PTM) sites, subcellular localization, signal peptide, transmembrane domain, secondary and tertiary structure followed by refinement and validations. In addition, B-cell epitopes were predicted and were screened using various web servers, while MHC-binding and CTL epitopes were predicted using IEDB and NetCTL servers, respectively. Results: The protein had 219 residues with a molecular weight of 25.55 kDa and alkaline isoelectric pH (7.5). This protein was stable, thermo-tolerant (aliphatic index: 78.04) and hydrophilic (GRAVY: -0.440). The predicted antigenicity scores were low and the protein was nonallergenic in nature. There were no transmembrane domain and signal peptide in the sequence. Moreover, several B-cell, MHC-binding and CTL epitopes were found in the EgGST protein, which could be further used in multi-epitope vaccines. Conclusion: Further studies are needed on the development of vaccines in vivo using EgGST alone or in combination with other antigens in the future.

The Role of Some Free-Ranging Animals in the Transmission of Multi-Host Species of Cryptosporidium Spp.

Background: We aimed to characterize Cryptosporidium spp. in rats, cats, pigeons, and crows. Methods: Fifty-five animal origin Cryptosporidium spp. genome were identified, genotyped and confirmed by nested PCR and of RFLP-PCR analysis as well as sequenced based on 18s rRNA and gp60 genes in Tehran (2012-2019). Finally, the phylogenetic analysis was performed by MEGA software (version 7). Results: By the molecular method, Cryptosporidium spp. were detected in 24 (15.2%), 15 (15%), 2 (2%) and 13 (13%) cases of wild rats, cat, pigeon, and crow, respectively. Among the identified species by the RFLP pattern, most isolates were identified as C. parvum (24/157) 17.8% in rats, (15/100) 15% in cats, (13/100) 13%in crew and (2/100) 2% in pigeons; and the rest of the cases were C. muris and C. felis. The results of sequencing did not prove the existence of C. parvum, C. felis, C. muris, and rat genotype. Subtyping of C. parvum was indicated that the dominant subtype family belongs to the IId family and the subtype A20G1 was the most common subtype detected in all hosts while A19G1 was detected in one isolate of cat and pigeon. Conclusion: Free-ranging animals are infected by species/subtype of Cryptosporidium, which can infect humans. This shows by itself the hygienic importance of the free-ranging animals in urban ecosystems. In the transmission of human cryptosporidiosis, the multi-host Cryptosporidium species such as C. parvum, C. felis, and C. muris can be transferred potentially from these animals to humans.

Assessment of tissue levels of miR-146a and proinflammatory cytokines in experimental cerebral toxoplasmosis following atovaquone and clindamycin treatment: An in vivo study.

BACKGROUND: Despite recent advances for treating cerebral toxoplasmosis (CT), monitoring the parasite burden and treatment response is still challenging. miRNAs are small non-coding RNAs with regulatory functions that can be used in diagnosis and treatment monitoring. We investigated the changes in miR-146a, BAG-1 gene, IL-6, and IL-10 tissue levels in the brain of BALB/c mice with chronic CT caused by the PRU strain of T. gondii following anti-parasitic and antibiotic treatment. METHOD: Fifty-three 6-to 8-week-old BALB/c mice were infected using intraperitoneal inoculation of cerebral cysts of T. gondii PRU strain and then divided into five groups as follows: group 1 included mice treated with 100 mg/kg/d Atovaquone (AT), group 2 included mice treated with 400 mg/kg/d clindamycin (CL), group 3 included mice treated with combination therapy (AT + CL), group 4 included infected untreated mice as a positive control (PC), and; group 5 included uninfected untreated mice as negative control (NC). After the completion of the treatment course, tissue level of mir-146a, miR-155, BAG-1 gene, IL-6, and IL-10 was investigated with real-time polymerase chain reaction. The IL-6/IL-10 ratio was calculated as an indicator of immune response. Moreover, brain cyst numbers were counted on autopsy samples. RESULTS: miR-146a, IL-6, IL-10, and BAG-1 genes were expressed in PC, but not in the NC group; miR-146a, IL-6, IL-10, and BAG-1 gene expression were significantly lower in AT, CL, and AT + CL compared with PC. MiR-146a and BAG-1 levels in AT and CL were not different statistically, however, they both had lower levels compared to AT + CL (P < 0.01). There was no difference in the expression of IL-6 and IL-10 between treatment groups. BAG-1 expression was significantly lower in AT, than in CL and AT + CL (P < 0.0089 and < 0.002, respectively). The PC group showed a higher ratio of IL-6/IL-10, although this increase was not statistically significant. It is noteworthy that the treatment with AT reduced this ratio; in the inter-group comparison, this ratio showed a decrease in the AT and AT + CL compared to the PC. The number of brain tissue cysts was significantly lower in AT, CL, and AT + CL, than in PC (p < 0.0001). AT had significantly lower brain cysts than CL and AT + CL (P < 0.0001). CONCLUSION: It seems that the factors studied in the current research (microRNA and cytokines) are a suitable index for evaluating the response to antiparasitic and antibiotic treatment. However, more studies should be conducted in the future to confirm our findings.

The Frequency of Intestinal Parasitic Infections in COVID-19 Patients: A Case-Control Study in Tehran, Capital of Iran.

The present study was done to evaluate the prevalence of intestinal parasitic infections (IPIs) in patients with COVID-19 in health care centers (Imam Reza and Golestan hospitals), Tehran, capital of Iran. By designing a matched case-control study, 200 fecal samples were collected for each of the COVID-19 patients and healthy individuals. Nasopharyngeal/oropharyngeal swab samples were collected from all participants for the diagnosis of COVID-19. RNA extraction was performed, and then real time reverse-transcription polymerase chain reaction (rRT-PCR) assay was applied to detect viral RNA. Considering the lung complications, 25%> lung complications was detected in 49 patients, 25-49% in 42 patients, and 50%≤ in 109 patients. Fecal samples were examined using different parasitological techniques. After nested-PCR, sequencing was applied to identify Cryptosporidium spp. and microsporidia spp. A relatively lower prevalence of IPIs was detected among control group (7.5%), than in COVID-19 patients (13%), though not significant (P=0.13). The most prevalent parasite among patients was Blastocystis sp. (6%). Also, 13.76% of IPIs were detected in inpatients with more than 50% lung complication. As well, a remarkably significant difference in IPIs was observed among diarrheic COVID-19 patients, in comparison with nondiarrheic patients (P < 0.00001). Moreover, the isolated sequences in the present study belonged to C. parvum subtype IIa and Enterocytozoon bieneusi genotypes D and Peru 8. In conclusion, more epidemiological and clinical research studies are needed to better understand the status and interaction of IPI in COVID-19 in Iran and other countries.

Thymoquinone Effect on Leishmania tropica/infantum and Leishmania-Infected Macrophages.

INTRODUCTION: Leishmania is a parasitic protozoan that tries to enter and amplify within macrophages. Macrophage cells are also immune defense cells that phagocyte many microbes like bacteria, fungi, as well as parasites like Leishmania spp. However, they are unable to kill this parasite that resides in the phagosomes of contaminated macrophages and multiplies in these macrophages, leading to the destruction of contaminated macrophages and the emerging of Leishmania wounds. A large number of current therapies for Leishmania cure have adverse effects, or parasites have developed resistance to some of these therapies, so a better therapy for the cure of Leishmania is required. Thymoquinone is one of the Nigella Sativa ingredients with numerous biological effects, such as antioxidant as well as antimicrobial effects on a variety of microbes, namely fungi, bacteria, as well as parasites like Leishmania spp. The impacts of Thymoquinone on Leishmania tropica and Leishmania infantum, as well as Leishmania-infected macrophages, were examined in this study. METHODS: The impact of various Thymoquinone dosages on L. tropica and L. infantum promastigotes and amastigotes was examined in vitro. Flow cytometry, as well as MTT, was also applied to examine the cytotoxic activity of Thymoquinone on promastigotes of L. tropica and L. infantum, as well as the incidence of apoptosis. The amastigote assay is also utilized to calculate the % of contaminated macrophages as well as the number of the present parasites in each macrophage. RESULTS: The percentage of macrophages contaminated with L. tropica and L. infantum amastigotes after medicating with 20 µM of Thymoquinone was 23% and 19%, respectively. Also, after medicating with 10 µM of Thymoquinone, these percentages were 32% and 31%, respectively. Flow cytometry indicated that Thymoquinone caused 33.9% and 31.4% apoptosis in L. tropica and L. infantum, respectively. As determined by the promastigote assay, the inhibitory concentration (IC50) of Thymoquinone for L. tropica and L. infantum was 9.49 µM and 12.66 µM, respectively. The results of the promastigote and amastigote assay show that with an increase in Thymoquinone doses, its ability to kill Leishmania parasites increases, too. CONCLUSION: According to the results of the study, Thymoquinone has a potentially lethal impact on L. tropica and L. infantum promastigotes as well as amastigotes (within leishmania contaminated macrophages).

Evaluating leishmanicidal effects of Lucilia sericata products in combination with Apis mellifera honey using an in vitro model.

Leishmaniasis is a zoonotic disease caused by an intracellular parasite from the genus Leishmania. Lack of safe and effective drugs has increasingly promoted researches into new drugs of natural origin to cure the disease. The study, therefore, aimed to investigate the anti-leishmanial effects of Lucilia sericata larval excretion/secretion (ES) in combination with Apis mellifera honey as a synergist on Leishmania major using an in vitro model. Various concentrations of honey and larval ES fractions were tested against promastigotes and intracellular amastigotes of L. major using macrophage J774A.1 cell line. The inhibitory effects and cytotoxicity of ES plus honey were evaluated using direct counting method and MTT assay. To assess the effects of larval ES plus honey on the amastigote form, the rate of macrophage infection and the number of amastigotes per infected macrophage cell were estimated. The 50% inhibitory concentration (IC50) values were 21.66 µg/ml, 43.25 60 µg/ml, 52.58 µg/ml, and 70.38 µg/ml for crude ES plus honey, ES >10 kDa plus honey, ES <10 kDa plus honey, and honey alone, respectively. The IC50 for positive control (glucantime) was 27.03 µg/ml. There was a significant difference between viability percentages of promastigotes exposed to different doses of applied treatments compared to the negative control (p≤ 0.0001). Microscopic examination of amastigote forms revealed that dosages applied at 150 to 300 µg/ml significantly reduced the rate of macrophage infection and the number of amastigotes per infected macrophage cell. Different doses of larval products plus honey did not show a significant toxic effect agaist macrophage J774 cells. The larval ES fractions of L. sericata in combination with A. mellifera honey acted synergistically against L. major.

The Larval Stages of Echinostoma spp. in Freshwater Snails as the First and Second Intermediate Hosts in Gilan and Mazandaran Provinces, Northern Iran.

Background: Identification of the larval stages of Echinostoma spp. in freshwater snails is an essential guide to continue monitoring the possibility of their transmission and the potential of echinostomiasis in areas where trematodes are the primary agent of parasitic diseases. The aim of this study was investigate Echinostoma using morphological and molecular techniques. Methods: The study was conducted in Gilan and Mazandaran Provinces, northern Iran, from April 2019 to October 2021. Overall, 5300 freshwater snails were randomly collected and were identified using external shell morphology. Meanwhile, snails infected with trematodes were studied via shedding and dissecting methods. Larvae stages of Echinostoma were identified and the genomic DNA of the samples was extracted. The PCR amplification of the ITSI gene was carried out for 17 isolates and products were sequenced. Seven sequences were deposited in GenBank. Results: Totally, 3.5% of snails containing three species (Stagnicola sp., Radix sp. and Planorbis sp.) were infected with two types of cercaria, E. revolutum with 37 and Echinostoma sp. with 45 spines in the collar. Moreover, 35% of the snails were infected with Echinostoma spp. metacercaria. Phylogenetic analysis illustrated that isolates were included in two ITSI haplogroups. Conclusion: Results showed the potential hazard of a zoonotic parasite as Echinostoma in northern Iran. The potential of disease environmental relationship investigation and resource control optimization is necessary for effective disease prevention and health management.

Trematode Cercariae from Lymnaea gedrosiana in the Caspian Sea Littoral in Iran: a one health concern.

Introduction: Lymnaea gedrosiana snails are hosts to a variety of trematode cercaria of public and veterinary health importance. In Guilan Province, Iran, a region with a high level of fish and bird farming and wetlands important for migratory birds, little is known about the trematode cercaria from L. gedrosiana. Methods: From April 2020 to October 2021, six freshwater sites in Guilan Province were sampled for Lymnaeidae snails three times per season (spring, summer, autumn and winter). Snails were exposed to light and heat to induce cercaria shedding and shredded cercaria were identified morphologically and molecularly. Results: In total, 5,712 Lymnaeidae snails were collected of which 3,288 (57.6%) were identified to be L. gedrosiana with 54.3% containing trematode cercaria. Snail and cercaria recovery were highest in the spring and summer. Trematode cercaria identified included Telorchis assula, Hypoderaeum conoideum, Apharyngostrigea pipientis, Sanguinicola cf. inermis, Opisthioglyphe ranae, Diplostomum pseudospathaceum, and Australapatemon burti. Discussion: The four trematodes D. pseudospathaceum, S. inermis, A. burti, and A. pipientis have not been previously reported in Iran; all four of these can infect migratory birds. The most common cercaria found, H. conoideum (18.3% of the snails) is of zoonotic importance. The third most common cercaria found, S. inermis (10.0% of the snails) is detrimental to fish production. Given the importance of the wetlands in the region for wildlife and migratory birds as well as the number of fish and bird farms in the area, efforts to control L. gedrosiana snails are needed to protect wildlife and human health. In addition, monitoring programs should be implemented to identify and prevent introductions of new trematode species.

Microscopic and Molecular Identification of Cyclospora cayetanensis and Cystoisospora belli in HIV-Infected People in Tabriz, Northwest of Iran.

Opportunistic pathogens such as Cryptosporidium, Cystoisospora belli, and Cyclospora cayetanensis cause various gastrointestinal and non-digestive disorders in people with HIV/AIDS. These symptoms are especially severe in HIV-infected people who have a CD4+ count of less than 200 cells/mL. This study aimed to determine the prevalence of C. belli and C. cayetanensis infections among people living with HIV in Tabriz, northwest of Iran. This descriptive study was performed on 137 people with HIV who had been referred to behavioral disease counseling centers in Tabriz. Then, after receiving written consent, fecal samples were collected and evaluated for the detection of parasitic infections using direct methods and modified acid fast staining, as well as polymerase chain reaction (PCR).From the 137 fecal samples collected (98 males and 39 females, between 20 and 40 years old), 1.5% were positive for C. cayetanensis and 2.9% were positive for C. belli. Due to the prevalence of C. cayetanensis and C. belli in people with HIV in Tabriz, essential measures, including personal hygiene training for infection control and prevention, seem necessary.

Anti-parasitic activity of a chimeric peptide Cecropin A (2-8)-Melittin (6-9) (CM11) against tachyzoites of Toxoplasma gondii and the BALB/c mouse model of acute toxoplasmosis.

Toxoplasmosis is a zoonotic disease that infects most animals, including humans. Pyrimethamine/sulfadiazine is the standard treatment for toxoplasmosis. Although this treatment has been successful, it is often associated with side effects that cannot be tolerated. Therefore, various compounds have been proposed as alternative treatments for toxoplasmosis. Antimicrobial peptides (AMPs) act on various pathogens, from viruses to protozoa. The purpose of the present study was to evaluate the effects of CM11 on in vitro and in vivo Toxoplasma gondii infection. For in vitro experiments, VERO cells were treated with different concentrations of CM11 (1-128 µg/ml) compared to sulfadiazine (SDZ) (0.78-100 µg/ml). MTT and lactate dehydrogenase (LDH) assays evaluated the cell viability and plasma membrane integrity. Then, the inhibitory concentration (IC50) values were determined for treating tachyzoites of T. gondii before or on cells previously infected. Annexin V-FITC/propidium iodide (PI) staining was used to distinguish viable and apoptotic cells. The effect of CM11, SDZ, and a combination of CM11 and SDZ was evaluated in the BALB/c mouse model of acute toxoplasmosis. CM11 was effective on tachyzoites of T. gondii and had a time and dose-dependent manner. The results of the MTT assay showed that the CC50 values of CM11 and SDZ were estimated at 17.4 µg/ml and 62.3 µg/ml after 24-h, respectively. The inhibitory concentration (IC50) of CM11 and SDZ on infected cells was estimated at 1.9 µg/ml and 1.4 µg/ml after 24-h, respectively. The highest rate of apoptosis (early and late) in high concentrations of SDZ and CM11 was determined for tachyzoites (2.13 % and 13.88 %), non-infected VERO cells (6.1 % and 19.76 %), and infected VERO cells (7.45 % and 29.9 %), respectively. Treating infected mice with CM11 and a combination of CM11 and SDZ had increased survival time. Based on the mentioned results, it can be concluded that CM11 has a beneficial effect on tachyzoites of T. gondii in vitro. The result of the mouse model suggests that CM11, either alone or in combination with other chemotherapeutic agents, could be a potential therapeutic for toxoplasmosis. Hence, antimicrobial peptides could be applied as promising anti-toxoplasma agents for treating toxoplasmosis.

Genotyping of Acanthamoeba Isolated from Hospital Environments and Thermal Water of Recreational Baths in Markazi Province, Iran.

Background: Due to the opportunism character of Acanthamoeba, the presence of this parasite in the thermal water of recreational baths and hospital environments can be a risk to the health of staff, patients and others. The aim of this study was to investigate the distribution of potentially pathogenic Acanthamoeba genotypes isolated from the hospital environment and the thermal water of recreational baths in Markazi Province, central Iran. Methods: Overall, 180 samples including thermal water from recreational baths in Mahallat City and dust, soil and water from different hospitals of Arak, Farahan and Komijan cities, central Iran were collected. The presence of Acanthamoeba was investigated using microscopic examination and molecular methods. The PCR and sequencing was performed based on a specific 18S fragment of ribosomal DNA. Results: Based on the microscopic survey, totally 134 positive samples were detected including 35% in thermal water samples and 44.7% in hospital samples. In molecular analysis, 53.5% of the samples were identified as Acanthamoeba and 46.7% as Protacanthamoeba bohemica. The genotypes were detected as T4 (33.3%), T2 (10%), T11 (6.7%), and T5 (3.3%). Conclusion: The T4 was the most common genotype found in hospitals sampling sites while the T2 genotype and P. bohemica were detected in thermal water sampling sites.

Investigation of antileishmanial activities of CaO nanoparticles on L. tropica and L. infantum parasites, in vitro.

Stibogluconate sodium and meglumine antimoniate are the main antimonials utilised as the primary treatment option for leishmaniasis. However, have a number of side effects that limit their use. Development of nanoparticles (NPs) use in biological research and remarkable antimicrobial effects and unique optical and structural properties of CaO NPs have motivated this study to evaluated the effect of different times/dilutions of CaO NPs on Leishmania tropica and Leishmania infantum. To evaluate the antileishmanial activity of CaO NPs, the cytotoxic effect of CaO NPs against L. tropica and L. infantum amastigotes, promastigotes, as well as macrophages, was evaluated using counting and MTT assay after adding different concentrations of CaO nanoparticles (800-6.25 µg/ml) to the parasite culture. The possible apoptosis by CaO NPs were evaluated via flow cytometry assay. The XRD-pattern related to CaO nanoparticles indicating the cubic phase structures. According the effects of nanoparticle on promastigotes the IC50 values of CaO nanoparticles within 72 h were 19.81 µg/ml for L. tropica and 22.57 µg/ml for L. infantum. The percentage of the normal, apoptotic, and necrotic cells was estimated to be 82.6%, 14.81%, and 2.69% for L. tropica, and 73.6%, 23.89%, and 2.58% for L. infantum, respectively. Our results showed acceptable in vitro activity level of CaO NPs against L. tropica and L. infantum promastigotes as well as intracellular amastigotes. CaO NPs were more effective against L. infantum compared to L. tropica in vitro study.

Frequency, subtypes distribution, and risk factors of Blastocystis spp. in COVID-19 patients in Tehran, capital of Iran: A case-control study.

Background: Recent theories on the possible interactions between the intestinal parasites and COVID-19 have stated that these co-infections may cause immune imbalance and further complications in the affected patients. Until now, there is no data about Blastocystis subtypes as an intestinal parasite in COVID-19 patients. Therefore, the present work was done to evaluate the molecular prevalence of Blastocystis spp. and related risk factors in Iranian patients with COVID-19. Method: Stool samples were gathered from 200 COVID-19 patients and 200 control, being matched regarding age, gender and residence. Then, stool samples were surveyed by parasitological methods, including direct slide smear and formalin-ether concentration. In the following, PCR and sequencing were used to detect Blastocystis spp. and their subtypes. Results: The frequency of Blastocystis spp. in patients with COVID-19 (7.5%; 15/200 by molecular method vs. 6%; 12/200 by microscopy method) was slightly higher than in individuals without COVID-19 (4.5%; 9/200 by molecular method vs. 4%; 8/200 by microscopy method), this difference was not statistically significant (P value = 0.57 for molecular method vs. P value = 0.81 for microscopy method). Regarding associated factors for Blastocystis spp., we found significant differences regarding the residence (rural), loose and watery stool with diarrhea, and duration of treatment (6 weeks <) in the COVID-19 group. Blastocystis ST3 was the most common subtype in the patients with COVID-19 and control group. Conclusions: Based on this results, health education, improved sanitation and good personal hygiene are highly recommended to prevent Blastocystis in COVID-19 patients.

In vitro and in vivo study on antiprotozoal activity of calcium oxide (CaO) and magnesium oxide (MgO) nanoparticles on promastigote and amastigote forms of Leishmania major.

BACKGROUND: Currently, anti-leishmanial drugs have been developed. However, the available compounds have several side effects such as drug resistance and toxicity that cause some limitation for use. The development of nanoparticles (NPs) use in biological research and the proven effectiveness of CaONPs and MgONPs on bacteria and fungi, along with the lack of information about its antileishmanial effects, have motivated this study. CaO and MgONPs possess considerable antibacterial effects because of their alkalinity and active oxygen species. This study has taken into account the impacts of these two NPs on the L. major in vitro and in vivo. METHODS: To evaluate the antileishmanial activity of NPs, the cytotoxic effect of CaONPs, MgONPs, and MgOCaONPs against L. major amastigotes, promastigotes, as well as macrophages, was evaluated using counting or MTT assay. The possible apoptosis of L. major by CaONPs, MgONPs, and MgOCaONPs was evaluated via flow cytometry assay. For in vivo study, BALB/c mice were allocated to five groups and the lesions of infected mice with L. major promastigotes were treated with a 200 µg/mL concentration CaONPs, MgONPs, and MgOCaONPs, then the mice underwent a 4-week follow-up to examine the wound diameter and survival rates. RESULTS: The XRD-pattern related to CaONPs and MgONPs indicating the cubic phase and Rocksalt cubic structures. According the effects of nanoparticle on promastigotes the IC50 values of CaONPs, MgONPs, and MgOCaONPs within 72 h were 7.9 ug/mL, 10.3 ug/mL, and 8.0 ug/mL respectively. CaONPs, MgONPs, and MgOCaONPs induced apoptosis in about 7.8%, 53.57%, and 12.8% of promastigotes. All mice presented lesions. MgONPs was the most effective in reducing the size of the lesions. CONCLUSION: According to the results of the present research, MgONPs and CaONPs showed good in vitro and in vivo effects on L. major promastigotes and intracellular amastigotes especially MgONPs, and also it seems that MgONPs are applicable in Leishmania infection treatment due to their potential antileishmanial effects.

Molecular characterization and genotyping of Toxoplasma gondii in free-living animals in Iran: Effect of One Health.

To understand the transmission of Toxoplasma gondii, this parasite's genetic diversity distribution in free-living hosts is essential. This research's objective is the molecular genotyping of T. gondii isolates from the brain and muscles of Columbidae, Corvidae, Rattus, and Felidae of Mianeh County, East-Azerbaijan Province, Northwest Iran. Three hundred fifty samples were taken. For the genotyping of T. gondii, the GRA6 gene was amplified and digested by the Tru1I (MseI) enzyme. Results of RFLP were confirmed by sequencing and phylogenetic analysis. In total, 52%, 34%, 24%, and 50% of Columbidae, Corvidae, Rattus, and Felidae were positive for T. gondii DNA, respectively. All isolated Columbidae were identified as genotype III (100%). Also, 94.1% and 5.9% of Corvidae isolates, 84.4% and 15.6% of the Rattus isolates, and 51.7% and 48.3% of the Felidae isolates belonged to genotypes III and II, respectively. This study is the first to evaluate genetic similarity and phylogenetic analysis between many definitive and intermediated hosts in northwestern Iran. The finding indicates that the T. gondii cycle is maintained among these hosts. As a result, their presence in the environment can be a risk factor for transmitting the infection to humans. Due to demographic and geographic differences in various regions, further studies are required to determine the genetic population structure.

Contamination of Vector Snails with the Larval Stages of Trematodes in Selected Areas in Northern Iran.

Background: Identification of freshwater snails and possible trematodes transmission sites are essential to continue monitoring the potential for disease outbreaks in areas with a history of parasitic infections. We aimed to search some areas in the margin of the Caspian Sea, northern Iran to identify the snail fauna of this area and verify the contamination of vector snails. Methods: More than 5,308 snails from 51 diverse and permanent habitats were studied from April 2019 to October 2021. Snails were collected randomly and identified using shell morphology. Trematode infection in snails was investigated by the release of cercariae and dissection methods. Results: Five families of freshwater snails including Lymnaeidae, Physidae, Planorbidae, Bithyniidae, and Viviparidae were investigated in the Caspian Sae Litoral of Iran. Physidae were found as the most prevalent snails (55.1%) followed by Lymnaeidae (29.4%). The parasitize rate was observed as 20% using releasing cercaria technique. Echinostomatoidea (31%), Schistosomatoidea (8%), and Diplostomoidea (21%), and Plagiorchioidea (40%) were seen as detected parasites. Meanwhile, 60% of the studied snails illustrated the other stages of trematodes. Conclusion: The rate of infection of snails with different cercaria in northern Iran is significant. It needs further deep studies to clarify the situation of zoonoses transmitted by snails in the region. Policy makers should pay attention more to this area in terms of monitoring the snail-transmitted diseases.