Global prevalence of Giardia duodenalis in cattle: A systematic review and meta-analysis.

Giardia duodenalis is an important intestinal parasite responsible for diarrhea in humans and animals worldwide. Up to now, G. duodenalis infections in cattle have been reported in many studies around the world. Hence, the aim of the present study is to report on the distribution of G. duodenalis in cattle at global scale and to evaluate the global prevalence, risk factors and genetic characterization of G. duodenalis infection among cattle worldwide. International databases were systematically searched to identify relevant studies. A random-effects meta-analysis model was used to estimate the overall and the subgroup-pooled prevalence of G. duodenalis across studies, and the variance between studies (heterogeneity) was quantified by I2 index. One hundred and fifty-eight articles (including 195 datasets), from 48 countries met eligibility criteria for analysis. Considering detection methods, the pooled prevalence was estimated to be 24% (95% confidence interval (CI), 19-30%) using copro-antigen techniques, 22% (95% CI, 17-28%) using molecular, and 16% (95% CI, 12-20%) using microscopic detection. Molecular methods showed that the highest number of reports were associated with assemblage E (45/46; 97.83% studies), assemblage A (33/46; 71.74% studies) and assemblage A+E (10/46; 21.74% studies). The pooled prevalence different of subgroups (WHO regions, countries, and type of cattle) were analyzed separately. Moreover, a significant association was observed between G. duodenalis infection with cattle suffering from diarrhea (odds ratio (OR), 2.61; 95% CI, 1.50-4.55) and pre-weaned calves (OR, 1.79; 95% CI, 1.08-2.95). These results suggest that the corresponding control scheme and effective management measures should be formulated to reduce the transmission of G. duodenalis infection according to the difference of geographical conditions in different areas.

Seroprevalence, risk factors, and clinical symptoms of Toxocara spp. infection among children 3-15 years old in northern Iran.

The World Health Organization has categorized toxocarosis as a neglected tropical disease despite its significant impact on high-risk groups such as children. This study aimed to investigate the seroprevalence, risk factors, and clinical symptoms of Toxocara spp. infection among children 3-15 years old in northern Iran. A total of 386 children were enrolled in the study. All serum samples were tested for the presence of IgG antibodies against Toxocara spp. infection using an enzyme-linked immunosorbent assay. Moreover, relevant risk factors and clinical symptom data were obtained using questionnaires. Data analysis was performed using the SPSS software version 24. The overall seroprevalence of Toxocara spp. infection was found 2.85 % (11/386). However, Toxocara spp. infection was high for some risk factors, including eating soil (14.3 %), contacting cats (6.7 %), and consuming raw vegetables (3.7 %). However, there were no statistically significant differences regarding the risk factors and socio-demographic characteristics. Considering the clinical symptoms, Toxocara spp. infection was different in children with eosinophilia (20 %), ocular disorders (8.3 %), skin disorders (7.7 %), liver disorders (4.5 %), and stomach ache (4.2 %), although not statistically significant. The results revealed that the seroprevalence of Toxocara spp. infection was relatively low in children in northern Iran. It is suggested to conduct more studies in different parts of Iran to gain a deeper understanding of the toxocarosis seroprevalence and its status in high-risk groups such as children with asthma, hypereosinophilic syndrome, allergic skin disorders, and epilepsy.

Serological Study of Fascioliasis Using Indirect ELISA in Gorgan City, Golestan Province, Northern Iran.

BACKGROUND: Fascioliasis is a neglected zoonotic disease, caused by Fasciola species in human and livestock. We aimed to detect the seroprevalence of human fascioliasis Gorgan City, Golestan Province, northern Iran using ELISA method in 2017. METHODS: Overall, 612 serum samples were analyzed. A relevant questionnaire for demographic data was obtained for all cases. An indirect ELISA test was used to detect IgG antibodies against Fasciola in the sera. The data analysis was performed employing SPSS program version 21. RESULTS: Eleven cases (1.79%) were seropositive for fascioliasis. The seroprevalence of fascioliasis was 1.9% and 1.1% among males and females, respectively. There was no statistically significant association between the fascioliasis and analyzed variables such as sex, age, residence, job, education, etc. CONCLUSION: This study was conducted only on the people referring to the Reference Laboratory of Gorgan. It cannot be distributed to the whole city. Thus, due to importance of the disease, finding the seroprevalence of fascioliasis in a comprehensive survey in Golestan Province should be accounted in further studies.

Seroprevalence of Hydatidosis in People Referring to Reference Laboratory of Gorgan, Golestan Province, Northern Iran 2017.

BACKGROUND: Hydatidosis is a neglected global zoonotic disease, caused by larval stage of the cestode Echinococcus granulosus in human and animal. Because of high economic and medical importance of the disease, this study was performed to find the seroprevalence of human hydatidosis in Gorgan City, Golestan Province, northern Iran. METHODS: In this cross-sectional study, blood samples were collected from people referring to Reference laboratory of Golestan University of Medical sciences in 2017. A relevant questionnaire was completed for demographic data for each person. Echinococcus IgG antibody was investigated by ELISA using native antigen B. The data were analyzed using SPSS software applying logistic regression. RESULTS: Overall, 612 blood samples were collected. Cut-off was considered 0.29. Sixteen cases (2.6%) were seropositive for hydatidosis. The seroprevalence of hydatidosis was 2.3% and 4.7% among males and females, respectively. There was no statistically significant correlation between the hydatidosis and investigated variables such as sex, age, tribes, residence, education, etc. CONCLUSION: The prevalence of human hydatidosis shows approximately the same range as other regions of Iran. Although due to the neighboring the Mazandaran Province reported as the highest seroprevalence of hydatidosis, we expected more rate of seropositivity.

Transient knockdown of Nucleoside transporter 4 gene expression as a therapeutic target in Leishmania major by antisense RNA: In vitro and in vivo studies.

BACKGROUND & OBJECTIVES: Leishmania parasites cause various clinical symptoms in humans such as cutaneous ulcers and fatal visceral diseases. These parasites cannot synthesize purine rings de novo and must uptake purines from their hosts via salvage. Salvage is regulated by permeases in the cell membrane. There are hundreds of membrane transporter proteins to receive nutrients in Leishmania. Nucleoside transporter 4 (NT4) is one of the purine transporters that is involved in enhancing the uptake of adenine in Leishmania major. They are important new drug targets for the treatment of leishmaniasis because they can be used to transport toxic purine analogs to kill parasitic cells, thus preventing the progression of the infection. The present study was conducted to silence the NT4 nucleobase involved in the salvage pathway to interrupt purine nucleotide membrane transport in the cells of L. major. METHODS: In this study, a 502 bp segment of NT4 gene sequence was selected and designed as antisense transcripts after insertion in the parasite. The NT4 construct was transfected into L. major promastigotes for in vitro study of gene expression. Then, BALB/c mice infected with transgenic Leishmania and wild-type strain along with the number and size of lesions were studied in vivo. RESULTS: The study showed that relative expression of NT4 gene in mutant Leishmania was lower than in the control on Day 3 to 20. The percentages and the number of amastigotes in infected macrophages with wild-type strain L. major were more than infected macrophages with mutant parasites. Infected BALB/c mice with transgenic Leish- mania showed a lower number and size of lesions than the BALB/c mice infected with wild-type strain. INTERPRETATION & CONCLUSION: The results of the study indicated that the use of antisense RNA reduced NT4 gene expression in L. major. Further, studies are needed to ascertain that the use of antisense can be considered as a new treatment for leishmaniasis.

Transient Down-Regulation of Nucleoside Transporter 3 Gene Expression as a Drug Target in Leishmania major Using Antisense RNA Technology.

BACKGROUND: This study was aimed to silencing the Nucleoside transporter 3 (NT3) permease nucleobases involved in the salvage pathway of Leishmania in order to disrupt purine nucleotide uptake in the parasite and consequently, destruction of the parasite. METHODS: Overall, 502 bp fragment of the NT3 gene sequence was designed to produce an antisense transcript upon entry of the vector into the parasite. The NT3 construct was transfected into L. major promastigotes and NT3 gene expression was studied in vivo and in vitro conditions. RESULTS: Relative expression NT3 gene in transgenic Leishmania was decreased in tenth day. The percentages and the number of amastigotes infected macrophages with transgenic L. major were less than infected macrophages with wild-type strain. Our results in two groups of BALB/c female mice (wild-type strain and mutant, n=4 each group) were showed that size and number of ulcers in BALB/c mice infected with transgenic Leishmania promastigotes were less than the BALB/c mice infected with wild-type parasites. CONCLUSION: The results indicate the use of antisense RNA reduces of NT3 gene expression in L. major. More studies are required to obtain a new approach for treating Leishmania infection.

First molecular report of Hydatigera krepkogorski (Schulz and Landa, 1934) in abdominal cavity of gerbil (Rhombomys opimus) in Iran.

A polycephalic larva of Taeniidae family isolated from abdominal cavity of a great gerbil (Rhombomys opimus) from Golestan province, northern Iran, was subjected to molecular analysis. Genomic DNA from the larva was obtained using a DNA extraction tissue kit. Polymerase chain reaction was performed for amplification of the partial 12S rRNA, cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase 1 (nad1) mitochondrial genes. BLAST analysis of DNA sequencing indicated 99.00% homology in 12S rRNA and cox1 genes and 98.00% homology in nad1 gene with Hydatigera krepkogorski (accession No. AB731762). The sequences of current isolate were deposited in GenBank by accession Nos. MF281971, MF281972 and MF281973 for 12 SrRNA, cox1 and nad1 genes, respectively. This study was the first report of molecular characterization of H. krepkogorski from Iran. Isolation and characterization of the adult stage from definitive host will help to better clarify incomplete life cycle and morphology data of this species in the world.

Differential Expression of Hox and Notch Genes in Larval and Adult Stages of Echinococcus granulosus.

This investigation aimed to evaluate the differential expression of HoxB7 and notch genes in different developmental stages of Echinococcus granulosus sensu stricto. The expression of HoxB7 gene was observed at all developmental stages. Nevertheless, significant fold differences in the expression level was documented in the juvenile worm with 3 or more proglottids, the germinal layer from infected sheep, and the adult worm from an experimentally infected dog. The notch gene was expressed at all developmental stages of E. granulosus; however, the fold difference was significantly increased at the microcysts in monophasic culture medium and the germinal layer of infected sheep in comparison with other stages. The findings demonstrated that the 2 aforementioned genes evaluated in the present study were differentially expressed at different developmental stages of the parasite and may contribute to some important biological processes of E. granulosus.

Comparison of ex vivo harvested and in vitro cultured materials from Echinococcus granulosus by measuring expression levels of five genes putatively involved in the development and maturation of adult worms.

Parts of the natural life cycle of Echinococcus granulosus can be retraced in vitro such as the development of protoscoleces into semiadult worms with three or more proglottids, or the redifferentiation of in vitro cultured protoscoleces into metacestode-like cystic structures. Most in vitro generated samples share-at the microscopical level-high similarities with those naturally grown, but developmental differences have also been documented, such as missing egg production in in vitro grown adults or unusual bladder/vesicle formation in protoscoleces cultured into the metacestode direction. The aim of the present study was to explore how far different in vitro generated stage-specific materials/structures match the natural situation on the transcriptome level, based on testing five exemplarily chosen different genes: the frizzled receptor eg-fz4 (posterior marker), the FGF receptor-like factor eg-fgfrl (anterior association), the cell differentiation protein eg-rcd1 (part of the CCR4-NOT complex, a key regulator of eukaryotic gene expression), the rapidly accelerated fibrosarcoma serin/threonin kinase eg-braf (part of the MAPK pathway involved, e.g., in EGF signaling) and the co-smad eg-smadD (downstream factor of TGFß/BMP2/activin signaling). These genes-tested via qPCR-were selected such as to allow a discussion on their potential role in the development of E. granulosus into the adult stage. Thus, testing took place with three ex vivo isolated samples, namely (i) egg-containing adult worms, (ii) invaginated protoscoleces, and (iii) protoscolex-free germinal layer tissue. Respective data were compared (a) with in vitro generated metacestode-like microcysts developed from protoscolices, and (b) different development stages of protoscoleces in vitro cultured toward adult maturation. As a finding, only eg-smadD and partially eg-fz4 showed high expression similarities between ex vivo harvested and in vitro cultured E. granulosus, thus suggesting a putative role in adult maturation. Conclusively, the fact of using "only" five genes did not allow answering the question if ex vivo and in vitro materials are similar on the transcriptome level. Another experimental restriction arises from different growth conditions of the in vitro cultured materials, and comparing these to the ex vivo harvested ones. Future experiments may solve the problems by using fully standardized E. granulosus sample collection and fully standardized culture conditions.

The role of GlcNAc-PI-de-N-acetylase gene by gene knockout through homologous recombination and its consequences on survival, growth and infectivity of Leishmania major in in vitro and in vivo conditions.

At present, there are no efficacious vaccines or effective drugs against leishmaniasis; therefore new and innovative control methods are urgently required. One way to achieve this important goal is through using reverse genetic engineering to evaluate important enzymes, proteins and macromolecules. One of the most important enzymes for Glycosylphosphatidylinositol (GPI) biosynthetic pathways is GlcNAc-PI-de-N-acetylase (GPI12). The molecular constructs were cloned in Escherichia coli strain Top 10 and confirmed by molecular methods and were transfected by electroporation into Leishmania major. We demonstrated that two alleles of the GPI12 gene in L. major were successfully removed and enabling the generation of a null mutant, which supports the idea that GPI12 is not an essential gene for the growth and survival of Leishmania and the homozygous knockouts of Leishmania are able to survive. We were able to produce a mutant parasite that caused no damaged to the host. Further investigations are essential to check the safety profile in laboratory animals.

Molecular cloning, expression and enzymatic assay of pteridine reductase 1 from Iranian lizard Leishmania.

BACKGROUND: Currently, there are no effective vaccines against leishmaniasis, and treatment using pentavalent antimonial drugs is occasionally effective and often toxic for patients. The PTR1 enzyme, which causes antifolate drug resistance in Leishmania parasites encoded by gene pteridine reductase 1 (ptr1). Since Leishmania lacks pteridine and folate metabolism, it cannot synthesize the pteridine moiety from guanine triphosphate. Therefore, it must produce pteridine using PTR1, an essential part of the salvage pathway that reduces oxidized pteridines. Thus, PTR1 is a good drug-target candidate for anti-Leishmania chemotherapy. The aim of this study was the cloning, expression, and enzymatic assay of the ptr1 gene from Iranian lizard Leishmania as a model for further studies on Leishmania. METHODS: Promastigote DNA was extracted from the Iranian lizard Leishmania, and the ptr1 gene was amplified using specific primers. The PCR product was cloned, transformed into Escherichia coli strain JM109, and expressed. The recombinant protein (PTR1 enzyme) was then purified and assayed. RESULTS: ptr1 gene was successfully amplified and cloned into expression vector. Recombinant protein (PTR1 enzyme) was purified using affinity chromatography and confirmed by Western-blot and dot blot using anti-Leishmania major PTR1 antibody and anti-T7 tag monoclonal antibody, respectively. The enzymatic assay was confirmed as PTR1 witch performed using 6-biopterin as a substrate and nicotinamide adenine dinucleotide phosphate as a coenzyme. CONCLUSION: Iranian lizard Leishmania ptr1 was expressed and enzymatic assay was performed successfully.