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Patients with diabetes are at an increased risk of intestinal parasitic infections (IPIs). We evaluated the pooled prevalence and OR of IPIs in patients with diabetes through a systematic review and meta-analysis. A systematic search was performed using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) protocol for studies reporting IPIs in patients with diabetes through 1 August 2022. The collected data were analyzed using comprehensive meta-analysis software version 2. Thirteen case-control studies and nine cross-sectional studies were included in this study. The overall prevalence of IPIs in patients with diabetes was calculated to be 24.4% (95% CI 18.8 to 31%). Considering the case-control design, the prevalence of IPIs in case (25.7%; 95% CI 18.4 to 34.5%) was higher than controls (15.5%; 95% CI 8.4 to 26.9%) and a significant correlation was observed (OR, 1.80; 95% CI 1.08 to 2.97%). Moreover, a significant correlation was seen in the prevalence of Cryptosporidium spp. (OR, 3.30%; 95% CI 1.86 to 5.86%), Blastocystis sp. (OR, 1.57%; 95% CI 1.11 to 2.22%) and hookworm (OR, 6.09%; 95% CI 1.11 to 33.41%) in the cases group. The present results revealed a higher prevalence of IPIs in patients with diabetes than in controls. Therefore, the results of this study suggest a proper health education program to preventing measures for the acquisition of IPIs in patients with diabetes., (© The Author(s) 2023. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Giardia duodenalis is a cryptic protozoan, which has eight assemblages (A-H). Assemblages A and B are the main genotypes reported from humans with probable anthroponotic and zoonotic transmission. The current study aimed to characterize G. duodenalis assemblages in tuberculosis (TB) patients and healthy subjects using multilocus genotyping (MLG). Thirty Giardia-positive stool samples, which were obtained from TB patients and healthy subjects were included in the study. After total DNA extraction, three β-giardin (bg), triosephosphate isomerase (tpi), glutamate dehydrogenase (gdh) genes were amplified and sequenced. Obtained sequences were compared to the GenBank database to characterize assemblages. Phylogenetic analysis using Maximum Likelihood (ML) and Tamura 3-parameter was performed for each gene. From 30 Giardia-positive subjects, 17 (57%) and 13 (43%) were from healthy and TB-infected subjects, respectively. There was no significant co-existence of Giardia and tuberculosis (P-value = 0.051). In addition, 14 (46.7%) and 16 (53.3%) of Giardia isolates were from asymptomatic and symptomatic subjects, respectively. PCR amplification was successful in 25 single samples (83.3%) consisted of 20 for tpi, 15 for bg, and 13 for gdh genes. Accordingly, 13/25 (52%) and 8/25 (32%) belonged to assemblage A and assemblages B, respectively, whereas 4/25 (16%) were either assemblage A or B with different genes at the same time. Significant correlation between assemblages and TB, age, and symptoms was not seen. The phylogenetic analyses represented no separation based on TB and gastrointestinal symptoms. Assemblage A was the predominant genotype in samples. The high frequency of assemblage AII indicated importance of anthroponotic transmission of Giardia in both healthy and TB patients. In addition, considering the exclusive reports of sub-assemblage AIII in wild ruminants, the presence of AIII in the current study have to be carefully interpreted. The inconsistency between the assemblage results of either bg or gdh loci with tpi gene signifies the insufficiency of single gene analysis and the necessity for MLG in molecular epidemiology of G. duodenalis., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Mohammad Rahimi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Dicrocoelium dendriticum is a broadly distributed zoonotic helminth, which is mainly reported from domesticated and wild ruminants. There is little data covering the molecular features of this trematode; therefore, current study aimed to molecularly analyze D. dendriticum in livestock., Methods: Totally, 23 samples of D. dendriticum were collected from cattle, sheep, and goat from Ilam, Lorestan, and Khuzestan, three west and south-west provinces of Iran from February to August 2018. After genomic DNA extraction, the internal transcribed spacer (ITS) 2 fragment was amplified and sequenced in samples. To investigate genetic variations through the ITS 2 fragment of obtained D. dendriticum, phylogenetic tree and network analysis were employed., Results: All 23 samples were successfully amplified and sequenced. Phylogenetic tree showed that our samples were clearly grouped in a clade together with reference sequences. There was no grouping based on either geographical regions or hosts. Network analysis confirmed the phylogenetic findings and showed the presence of nine distinct haplotypes, while our samples together most of sequences, which were previously submitted to the GenBank, were grouped in the Hap1., Conclusions: Our findings indicated that although ITS 2 fragment discriminate D. dendriticum, this fragment is not suitable to study intra-species genetic variations. Therefore, exploring and describing new genetic markers could be more appropriate to provide new data about the genetic distribution of this trematode., (© 2022. The Author(s).)

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 I 2 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., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Free-living amoebas (FLAs) can cause neurological and ocular complications in humans. Water supplies play a critical role in transmitting FLAs to humans. The aim of the present study was to investigate the presence of FLAs in various aquatic sources including drinking water, stagnant water, and surface water in Alborz province, northern Iran, using morphological and molecular techniques. A total of 70 water samples were collected from 34 drinking waters, 23 surface waters, and 13 stagnant waters. Filtration and cultivation were employed to isolate FLAs. PCR assay was applied by using the genus-specific primers on positive samples. Pathogenicity tests (osmo- and thermo-tolerance properties) were performed for Acanthamoeba spp., positive sample. Considering the morphological criteria, four positive samples of Acanthamoeba sp., three Vermamoeba sp., two mixed Vermamoeba sp. with Vahlkamfiids, and one mixed Acanthamoeba sp. with Vahlkamfiids were isolated. Five Acanthamoeba sp. isolates were amplified using the JDP primer pairs. Among them, two genotypes, T4 (three isolates) and T5 (two isolates) corresponding to A. lenticulata, were identified. Four V. vermiformis samples were confirmed using the sequencing. This study highlighted the occurrence of potentially pathogenic waterborne FLAs in water habitats associated with high human activity. The results of such research on the prevalence of FLAs, as a human hazard, should be communicated to health policymakers.

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., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (Copyright © 2022 Mirabedini et al. Published by Tehran University of Medical Sciences.)

Background: Microsporidiosis as a zoonotic disease has caused serious health problems in high-risk groups, including immunosuppressed individuals. Among the potential animal reservoirs of microsporidia, rodents play a key role due to close-contact with humans and their dispersion in different environments. Therefore, this systematic review and meta-analysis aimed to assess the global status and genetic diversity of microsporidia infection in different rodents., Methods: The standard protocol of preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines were followed. Scopus, PubMed, Web of Science, and Google Scholar were searched from 1 January 2000 to 15 April 2021. All peer-reviewed original research articles describing the molecular prevalence of microsporidia infection in rodents were included. Inclusion and exclusion criteria were applied. The point estimates and 95% confidence intervals were calculated using a random-effects model. The variance between studies (heterogeneity) were quantified by I 2 index., Results: Of 1695 retrieved studies, 22 articles (including 34 datasets) were included for final meta-analysis. The pooled global molecular prevalence (95% CI) of microsporidia infection in rodents was 14.2% (95% CI 10.9-18.3%). The highest prevalence of microsporidia was found in Apodemus spp. 27.3% (95% CI 15-44.5%). Enterocytozoon bieneusi was the most common pathogen (26/34; 76.47% studies) according to PCR-based methods, and the genotype D as the highest reported genotype (15 studies)., Conclusions: The findings of the study showed a relatively high prevalence of microsporidia infection in rodents as a potential animal reservoir for infecting human. Given the relatively high incidence of microsporidiosis, designing strategies for control, and prevention of microsporidia infection in rodents should be recommended., (© 2021. Witold Stefański Institute of Parasitology, Polish Academy of Sciences.)

Aim: This systematic review and meta-analysis evaluated the subtyped Blastocystis sp. isolated from humans in Iran., Background: Blastocystis sp. is an anaerobic intestinal protozoan that infects humans as well as domestic and wild animals, i.e. mammals, amphibians, reptiles, and arthropods., Methods: A comprehensive search for papers published before April 2022 was undertaken utilizing English and Persian databases. The following MeSH keywords were used in the electronic search: ( Blastocystis sp.) AND (molecular OR subtype) AND (prevalence OR epidemiology) AND Iran. The quality of the included studies was evaluated. Thereafter, a random-effects meta-analysis was conducted to estimate the pooled prevalence and odds ratios regarding the included studies., Results: A total of 32 studies comprised of five case-control studies and 27 cross-sectional studies met the eligibility criteria. The overall pooled prevalence of subtyped Blastocystis sp. in Iran was estimated to be 10% (95% confidence interval: 6 to 15%). Eight subtypes of Blastocystis sp. (ST1- ST7 and ST9) were identified in our study, of which ST3 was the most common subtype (0.04); 0.02-0.07). The difference in subtypes between two case and control groups in reported studies was not significant, but the odds ratio of infection by ST3 (0.98; 95% CI, 0.30 to 3.20) was higher in cases., Conclusion: The current systematic review showed that with the exception of ST8 and ST12, all human Blastocystis sp. subtypes reported in the world are found in different parts of Iran., Competing Interests: The authors declare no conflict of interest.

Objective: Heart transplant is one of the accepted treatments for some patients with advanced heart failure. Of note, transplant surgeries may cause different infections and complications for patients during the post-transplant period. A wide variety of opportunistic organisms caused these infections including bacteria, fungi, viruses, and protozoa particularly Free-living amoebae (FLA). This study aims to study the presence of pathogenic FLA from the oral cavity of post-heart transplant recipients., Methods: Throat swabs were collected from 80 patients who underwent post-heart transplant surgery. All swabs were immediately cultured in non-nutrient agar (2%). PCR and sequencing of 18S rRNA gene (DF3 region) of Acanthamoeba isolates were performed using genus-specific primers. Genetic associations among sequenced genotypes inferred by the 18S rRNA gene obtained by MEGA X and a phylogenetic tree were constructed using the maximum likelihood algorithm and Kimura 2-parameter model., Results: Out of 80 samples collected from post-heart transplant patients, six (7.5%) samples showed positive outgrowth of Acanthamoeba based on the page key and sequencing of the DF3 region. Sequence similarity of ASA1 by basic local alignment search tool(n) showed that five isolates (ANHT1, ANHT2, ANHT3, ANHT4, and ANHT5) belonged to Acanthamoeba T5 genotype corresponding to A. lenticulata and one strain (ANHT6) belonged to the T4 genotype., Conclusion: To the best of our knowledge for the first time, a comprehensive study of Acanthamoeba genotypes isolated from throat samples of heart transplant recipients is described. Heart transplantation patients can be colonized by FLA and are therefore at risk of developing an invasive infection. Physicians' awareness of central nervous system infections related to FLAs and preventive and control measures of patients with compromised immune status due to heart transplant surgery are of utmost importance., (© 2021 Wiley Periodicals LLC.)

Free-living amoeba (FLA) are ubiquitously distributed in the environment. However, they are also the causative agents of opportunistic infections in humans and other animals. A biofilm comprises any syntrophic consortium of microorganisms in which cells stick to each other and often also to a surface. Moreover, FLA have been detected in various biofilms around the world. Therefore, the present study aimed to check for presence of FLA in samples from household biofilms in Iran and to characterize them at the molecular level. A total of 69 biofilm samples collected from showerheads, kitchen areas, and bathroom sinks were analyzed. Positive samples for FLA were characterized at the morphological and molecular levels. Furthermore, the results of morphology analysis indicated that 26.08% (18/69) of biofilm samples were positive for Acanthamoeba spp., Vermamoeba genus, and Vahlkampfiids. According to sequence analysis, five strains of Acanthamoeba isolates related to the T4 genotype and two strains belonged to the T2 genotype. In addition, the pathogenic potential of Acanthamoeba -positive isolates was conducted using the tolerance ability test. The results of BLASTn of Vermamoeba sequences were similar to what was expected for Vermamoeba vermiformis . The above-mentioned reasons revealed that the relative high contamination of household biofilm samples with FLA may pose a risk for people using soft contact lenses and for patients with traumatic cataract. Our finding proposes that filtration should be performed in shower heads and indicates the need to monitor people at increased risk of Acanthamoeba keratitis.

Background: Microsporidia is a zoonotic pathogen with health consequences in immunocompromised patients. Small ruminants are a potential reservoir of microsporidia for humans in their vicinity. Hence, we aimed to evaluate the molecular prevalence of microsporidian infections with emphasis on Enterocytozoon bieneusi genotypes among sheep and goats at a global scale through systematic review and meta-analysis approach., Methods: The standard protocol of preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines were followed. Eligible prevalence studies on small ruminant microsporidiosis, published from 1 January 2000 until 15 April 2021 were gathered using systematic literature search in PubMed, Scopus, Web of Science and Google Scholar databases. Inclusion and exclusion criteria were applied. The point estimates and 95% confidence intervals were calculated using a random-effects model. The variance between studies (heterogeneity) was quantified by I 2 index., Results: In total, 25 articles (including 34 datasets) were included for final meta-analysis. The pooled molecular prevalence of microsporidia in sheep and goats was estimated to be 17.4% (95% CI: 11.8-25%) and 16% (95% CI: 11.2-22.4%), respectively. Likewise, the overall prevalence of E. bieneusi was estimated to be 17.4% (95% CI: 11.8-25%) for sheep and 16.3% (95% CI: 11.3-22.8%) for goats. According to internal transcribed spacer (ITS) gene analysis, E. bieneusi with genotypes BEB6 (15 studies) and COS-1 (nine studies) in sheep, and CHG3 (six studies) and BEB6 (five studies) in goats were the highest reported genotypes., Conclusion: The present results highlight the role of sheep and goats as reservoir hosts for human-infecting microsporidia. Therefore, this global estimate could be beneficial on preventive and control measures., (© 2021. The Author(s).)

Background: Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease. Its etiology remains largely unknown, although frequent concomitant inflammatory bowel disease (IBD) hints towards common factors underlying intestinal and bile duct inflammation. Herein, we aimed to explore the relative abundance of fecal microbiota in PSC-IBD patients compared to IBD-only subjects and controls., Methods and Results: We included 14 PSC-IBD patients, 12 IBD-only patients, and 8 healthy controls (HCs). A quantitative real-time PCR (qPCR) assay was used to determine a selection of bacterial phyla, families, and genera. Relative abundance of taxa showed that Bacteroidetes was the most abundant phylum among the patients with PSC-IBD (29.46%) and also HCs (39.34%), whereas the bacterial species belonging to the phylum Firmicutes were the most frequent group in IBD-only subjects (37.61%). The relative abundance of the Enterobacteriaceae family in fecal samples of PSC-IBD patients was similar to those with IBD-only, which was significantly higher than HCs (p value = 0.031), and thus, could be used as a PSC-IBD or IBD-only associated microbial signature., Conclusions: Our findings showed that intestinal microbiota composition in PSC-IBD patients was completely different from that of IBD-only patients. Further studies using large-scale cohorts should be performed to better describe the contribution of the gut microbiota to PSC pathogenesis with underlying IBD., (© 2021. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: Regarding the controversial role of Blastocystis in inflammatory bowel diseases (IBD) patients, it seems that this protozoan may lead to an overgrowth of some non-beneficial bacteria. The current study aimed to investigate the co-existence of Blastocystis and Clostridioides difficile in IBD patients., Methods: Stool samples of 102 IBD patients were collected and cultivated for C. difficile and Blastocystis. DNA extraction was performed on positive samples and C. difficile and Blastocystis were toxinotyped and subtyped, respectively. Fisher's exact test and logistic regression were employed to calculate the correlation between the existence of Blastocystis and its subtypes (ST) with C. difficile and its type of toxins. Also, the co-existence of Blastocystis and C. difficile with the frequency of defecations was evaluated., Results: Blastocystis and C. difficile were observed in 17 (16.7%) and 26 (25.5%) of stool samples, respectively. From 26 C. difficilepositive isolates, 24 (92.3%) and 2 (7.7%) were tcdA+/B+ and tcdA+/B-, respectively. Also, 10 (58.8%) and 7 (41.2%) were Blastocystis ST1 and ST3, respectively. Statistically significant correlations between co-existence of Blastocystis and C. difficile and co-existence of these microorganisms and frequency of defecation (P < .035) were seen. There was no statistically significant correlation between subtypes of Blastocystis and colonization of C. difficile or its toxinotypes., Conclusion: The co-existence of Blastocystis and C. difficile in IBD patients was observed in the current study. Moreover, it can be proposed that these microorganisms may have synergistic effects on their colonization in the gastrointestinal tract.

Free-living amoebae (FLA) including Acanthamoeba spp., Naegleria fowleri, and Balamuthia mandrillaris are among the many waterborne parasites. There is not much known about the possible FLA contamination of the irrigation resources watering crops and agricultural lands in this country. Forty-five water samples were collected from water canals used for irrigation of farmlands and tested for the presence of pathogenic FLA using morphological and molecular-based methodology. The ITS region of Naegleria species was sequenced and a phylogenetic tree was generated to confirm the genetic associations and taxonomic status of the identified Naegleria spp. A total of seven (15.5%) samples were positive for FLA. Molecular analyses identified two strains as N. philippinensis (isolates: PW1 and PW2) isolated from irrigation treated water. One strain showed high homology with N. americana (isolate: PW3). To the best of our knowledge, the present study is the first study to isolate the two strains of N. philippinensis and N. americana from agricultural waters in Iran. The presence of the FLA in irrigation waters should be considered a potential health threat for exposed farmers and other people.

Cryptosporidium and Giardia are two major protozoa reported from vegetables and environment. The prevalence of these parasites supposes to be different regarding the climate zones. This review aimed to evaluate the prevalence of Cryptosporidium and Giardia in vegetables according to the major climate zones in Iran. The results showed pooled prevalence 7% (95% CI: 2%, 14%) and 4% (95% CI: 3%, 6%) for Cryptosporidium spp., and Giardia spp., respectively. The prevalence of Giardia spp. in mountain, desert and semi-desert, and Mediterranean regions was 4% (95% CI: 2%, 6%), 5% (95% CI: 3%, 8%) and 7% (95% CI: 1%, 18%), respectively. Cryptosporidium spp. was reported 8% (95% CI: 0%, 65%), 6% (95% CI: 0%, 18%) and 4% (95% CI: 0%, 77%) from mountain, desert and semi-desert, and Mediterranean climate zones, respectively. This review suggests the higher prevalence of Giardia and Cryptosporidium in Mediterranean and mountain regions, respectively., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© Springer Nature Switzerland AG 2020.)

Fasciolosis is a public health problem originally transmitted from livestock. Although molecular analysis of the nuclear and mitochondrial genes allow discrimination between the two known species of Fasciola, F. hepatica and F. gigantica, these markers do not permit the detection of hybrid forms. On the other hand, molecular analysis of the pepck and pold genes from Fasciola do permit the detection of hybrid isolates and this study has not yet been performed on specimens from human and domesticated animals in Iran. Therefore, the aim of this study was to molecularly analyze Fasciola isolates using both conventional and the two new genetic markers mentioned above. Fifty-three adult Fasciola worms were isolated from the livers of 28 domesticated animals, including sheep, cattle and goat. Moreover, an adult worm was obtained from the bile duct of an infected woman during endoscopic retrograde cholangio-pancratography (ERCP). Targeted fragments from the ITS-1, NDI and COX1 genes were amplified using specific primers. PCR products were sequenced and genetically analyzed using MEGA v.7 and DnaSP software. Additionally, the pepck and pold regions were amplified and analyzed using multiplex PCR and RLFP-PCR, respectively. Multiple alignment of sequenced fragments showed highest similarity among the ITS-1 sequences isolated from all four hosts in comparison to the other genes. Furthermore, sequence diversity across the COX1 was higher than the NDI and ITS-1. Diversity among sequences isolated from cattle was higher than those from sheep and goat in all three genes. Tajima's D and Fu's Fs were negative and statistically significant for all the genes except ITS-1. Phylogenetic trees showed that the human F. hepatica isolate was closer to sheep isolates. The results of the pepck and pold analyses showed that all isolates were F. hepatica and there were no hybrid forms among samples. The molecular analyses corroborated this finding., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Microsporidia is a group of spore-forming microorganisms with zoonotic potential. This study aimed to compare intestinal microsporidia infections in cat owners and non-pet owners. In total, 210 fecal samples were collected from indoor cats, cat owners, and non-pet owners. DNA extraction was performed and the small subunit ribosomal RNA (SSU rRNA) gene was amplified. To characterize the genotypes, the internal transcribed spacer (ITS) fragment was amplified and sequenced. The phylogenetic trees were drawn to evaluate the relationship among Enterocytozoon bieneusi isolates. Two (2.9%) and one (1.4%) fecal samples from cat owners and one (1.4%) and two (2.9%) fecal samples from non-pet owners were positive for E. bieneusi and Encephalitozoon intestinalis, respectively. E. bieneusi was detected in two cat samples (2.9%). Same infection was not seen between infected cats and their owners. There was no significant difference between the prevalence rate of microsporidia among the cat owners and non-pet owners. Indeed, the genotypes L and type IV were seen in cats, while the genotype D was only detected in human. In this study, E. bieneusi and E. intestinalis were more prevalent among the cat owners and non-pet owners, respectively. Indeed, the higher prevalence of E. bieneusi in cats and their owners might be resulted from the worldwide distribution of this species.

Cryptosporidiosis, caused by the protozoan parasite Cryptosporidium spp., is an important zoonotic disease and is considered a global public health concern. Dogs are suggested as one of potential reservoirs for transmitting the Cryptosporidium infection to humans. However, there is a paucity of information about global patterns of occurrence of Cryptosporidium in dogs. A systematic review and meta-analysis were carried out to evaluate the global prevalence of Cryptosporidium infection among dogs. In this study, PubMed, Scopus, Web of Science and Google Scholar databases were systematically searched for relevant studies up until October 30, 2019. Finally, 127 articles (including 160 datasets) were eligible for inclusion in the systematic review and meta-analysis. The overall prevalence of Cryptosporidium infection was estimated at 8% (95 % CI: 5-11 %) using microscopic methods, 7% (95 % CI: 4-10%) using coprogantigenic methods and 6% (95 % CI: 4-9%) using molecular diagnostic methods. Molecular methods revealed that dogs were most frequently infected by C. canis (3.64 %) and C. parvum (1.28 %). The pooled prevalence different of subgroups (WHO regions, geographic and climate parameters, diagnostic methods, type of dog) were analyzed separately. The pooled odds ratio (OR) of Cryptosporidium was significantly higher than one for diarrhea status, with dogs suffering from diarrhea having a higher likelihood of Cryptosporidium infection, compared to dogs without diarrhea (OR; 3.61 95 % CI: 1.89-6.90%). The present study is the first systematic review and meta-analysis providing a comprehensive view of the global prevalence of Cryptosporidium in dogs and its related risk factors. Awareness of Cryptosporidium prevalence, risk factors, and disease complications for the health authorities, physicians, veterinarians and dog's owners is important for developing effective strategies to prevent infection., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Microsporidia are known opportunistic microorganisms and usually transmitted via the fecal-oral route. However, there is no information about human-infecting microsporidia in wildlife in Iran. This study aimed to investigate and analyze human-infecting microsporidia isolated from raccoons in north of Iran. Totally, 30 fecal samples were collected; then, DNA extraction was performed and specific fragments of the SSU rRNA gene of Enterocytozoon bieneusi and Encephalitozoon species were amplified. After amplification and sequencing the ITS, the results were compared to the GenBank database. Phylogenetic trees and network analysis were employed to explore probable relationships. E. bieneusi was the only detected microsporidia among samples. Genotyping showed the genotypes D, E, and RA in 15/18 (83.33%), 1/18 (5.55%), and 2/18 (11.11%) of samples, respectively. Novel genotypes RA1 and RA2 grouped together and apart from other genotypes. E. bieneusi genotypes D and E clustered with the genotypes previously reported from animals, humans, and environmental samples. Network analysis revealed six distinct sequence types among raccoon's isolates. This study demonstrated that E. bieneusi genotype D was the most prevalent microsporidia among raccoons. It seems that wildlife may play a role in dispersion of microsporidia spores., (© 2020 International Society of Protistologists.)

Although the main cause of appendicitis is unclear, infection with Enterobius vermicularis is suggested as a neglected risk factor. Since, there is no comprehensive analysis to estimate the prevalence of E. vermicularis in appendicitis; therefore, we conducted a global-scale systematic review and meta-analysis study to estimate the prevalence of E. vermicularis infection in appendicitis cases. PubMed, Scopus, Web of Science and Google Scholar databases were systematically searched for relevant studies published until 15 August 2019. Pooled prevalence of E. vermicularis infection was estimated using the random effects model. Data were classified based on the continents and countries. Moreover, subgroup analyses regarding the gender, the human development index (HDI), and income level of countries were also performed. Fifty-nine studies involving 103195 appendix tissue samples belonging to the individuals of appendicitis were included. The pooled prevalence of E. vermicularis infection was (4%, 95%CI, 2-6%), with the highest prevalence (8%, 95% CI: 0-36%) and lowest prevalence (2%, 95% CI: 1-4%) in Africa and Americas continents, respectively. With respect to countries, the lowest and highest prevalence rates were reported from Venezuela (<1%, 95% CI: 0-1%) and Nigeria (33%, 95% CI: 17-52%), respectively. Indeed, a higher prevalence was observed in females, as well as in countries with lower levels of income and HDI. Our findings indicate the relatively high burden of E. vermicularis infection in appendicitis cases. However, our findings suggest the great need for more epidemiological studies to depth understand overlaps between E. vermicularis infection and appendicitis in countries with lower HDI and income levels., Competing Interests: The authors have declared that no competing interests exist.

Purpose: Free-living amoeba (FLA) including Acanthamoeba spp., Balamuthia mandrillaris, and Naegleria are among the soil-born parasites. There are reports of FLA-related keratitis with a history of contact with soil and dust sources, particularly among the farmers. Due to lack of the previous studies on the farmland soils and a limited number of researches conducted on recreational soils in Iran, the present study was conducted., Methods: A total of 93 soil samples including farming lands and recreational places were tested for the presence of Acanthamoeba spp. Balamuthia mandrillaris, Naegleria, and Vermamoeba using morphological key and sequencing-based tools. Pathogenicity of Acanthamoeba positive strains was also evaluated. To verify genetic associations and taxonomic status of isolated amoeba, a phylogenetic tree was built by MEGA 5.05 software inferred by the 18S rRNA gene based on maximum likelihood algorithm., Results: Overall, 28 samples (30%) were contaminated with potentially pathogenic FLA, and according to the sequencing data, 17 strains were successfully sequenced. The isolated Acanthamoeba belonged to T2, T4, T5, mixed T4 and T5 contaminations, and T11. ITS sequencing revealed the occurrence of one strain of Naegleria canariensis. Four strains of Vermamoeba vermiformis were also confirmed. Morphological survey and PCR assay failed to show any positive results for Balamuthia mandrillaris. Pathogenic potential of the Acanthamoeba strains showed that T2, T4, and T11 genotypes were highly pathogenic, whereas T5 genotypes demonstrated lower pathogenic potential., Conclusion: The results indicate that soil could be a serious hazard to human health, and therefore, further studies are expected to investigate the source of infection in patients developing FLA-related diseases. The present study is the first to investigate FLA in the farmland soils in Iran and the first to report the presence of N. canariensis in the country.

Treated wastewater samples were collected, filtered using sterile 47-mm cellulose nitrate membrane and DNA extracted from the filtered materials. The presence of Blastocystis sp. was confirmed via polymerase chain reaction (PCR) targeting the SSU rRNA gene of Blastocystis sp. in 5/12 of samples. Based on the subtype analysis after sequencing, 2, 2 and 1 of ST2, ST6 and ST8 were detected among the isolates, respectively. Furthermore, both ST6s were allele 139, alleles 11 and 138 were identified in ST2 and the only ST8 was allele 95. The phylogenetic tree showed that one of ST2 was clustered together with those ST2 that were already reported from humans and animals. The presence of Blastocystis sp. in treated wastewater can indicate the potential role of this type of water for irrigation in the transmission of pathogenic microorganisms to downstream farmlands.

Aim: This study aimed to investigate the presence of oocyst of Cryptosporidium sp., and egg of soil-transmitted helminths in market vegetables in the north of Iran., Background: Fecal-oral transmission via consumption of contaminated food is the main route of transmission of intestinal parasites. Concerning the high risk of contamination of vegetable with intestinal parasites, raw consumption of crops can enhance the chance of transmission of intestinal parasites., Methods: In this study, we collected 34 pre-washed vegetable samples including spinach, mint, parsley, oregano, chives, savory, radish, coriander, basil and tarragon from local markets in Tonekabon City, North of Iran. All vegetable samples were washed using sterile PBS. Parasitological examinations, including direct examination and staining with Lugol's iodine and modified Ziehl-Neelsen were performed on the pellet resulted from the washing process., Results: The findings showed that 14/34 (41.17%) of collected samples were contaminated with at least one parasite. Eggs of Toxocara sp., Ascaris sp., Fasciola sp., Toxoascaris leonine , Trichuris sp. and Enterobius together with free-living larvae, amoeba cyst, cyst of Entamoeba coli and oocyst of Cryptosporidium sp., were observed among the positive samples. Furthermore, statistical analysis indicated that there was no significant correlation between parasitic contamination of vegetables and seasonal changes., Conclusion: This study signifies that some parasites due to their resistant cell wall usually remain in an environment with the harsh condition and thus, consumption of raw vegetables increases the risk of transmission of them., Competing Interests: The authors declare that they have no conflict of interest., (©2019 RIGLD, Research Institute for Gastroenterology and Liver Diseases.)

Background: Blastocystis and tuberculosis are two public health issues that are frequently reported in regions with low level of hygiene. Therefore, the current study aimed to investigate Blastocystis subtype and allele distribution in TB patients., Methods: Totally, 161 stool samples were taken from TB patients who were undergoing anti-MTB treatment. Stool samples were concentrated using conventional formalin-ether technique and examined using Lugol's iodine staining under light microscopy. DNA extraction was carried out and discriminative fragment was amplified and sequenced. With comparison in GenBank database, relevant subtypes and alleles were characterized and phylogenetically analyzed using MEGA v.7 and Tamura 3-parameter model., Results: In total, from 161 stool samples, 19 samples were suspected to be Blastocystis-positive. The expected fragment was amplified in 13 (8.07%) of samples. Accordingly, 11/13 (84.62%) of Blastocystis cases settled in urban and 2/13 (15.38%) were villagers. Close-contact with animals was also seen among 7/13 (53.84%) of samples. Subtype 1 (7/13; 53.84%) was the most prevalent followed by subtype 2 (5/13; 38.46%) and subtype 3 (1/13, 7.69%). All ST1 were allele 4, while alleles 9, 11 and 12 were seen in ST2 and allele 34 was the only allele observed in ST3. All three subtypes were clearly separated, while there was no separation between sequences from TB and non-TB patients., Conclusion: Blastocystis ST1 was the most prevalent subtype in TB patients and there was no difference between Blastocystis isolates from TB and non-TB human subjects., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Aim: To investigate the presence of intestinal parasites in tuberculosis patients who suffered from immunodeficiency disorders., Background: Tuberculosis is an important infectious disease that is endemic in some regions of Iran. However, there is a coverage in the endemicity areas of this infection with intestinal parasites., Methods: Stool samples were collected from 50 immunocompromised tuberculosis patients. Direct smear using the normal saline (0.85% NaCl solution) and Lugol's iodine staining were performed to detect trophozoite of parasites. Moreover, stool samples were concentrated using routine formalin-ether to detect protozoan cysts and helminth's ova/larvae. Specific staining techniques including Trichrome, Modified Ziehl-Neelsen and chromotrope 2R were employed to detect amoeba, Giardia spp., coccidian parasites and microsporidia., Results: From 50 participants, 42 (84%) and 8 (16%) were male and female, respectively. The mean age + SD of patients was 47.88 + 10.88 years. Among the participated patients, HIV positive, cancer, organ transplant and receiving corticosteroids were seen in 13, 10, 15 and 12 subjects, respectively. The prevalence of Intestinal parasites was 34 %( 17/50). Blastocystis (18%; 9/50), and intestinal helminth ( Enterobius vermicularis ) (2%; 1/50) were the most prevalent and less prevalent parasites, respectively. Statistical significance difference was not seen between presence of intestinal parasites and type of immunodeficiency., Conclusion: Our findings showed the high prevalence of intestinal parasites with majority of Blastocystis . Indeed, this study suggested that due to complicated immune conditions of TB patients with immunodeficiency disorders, this group of patients are at higher risk of infection by intestinal parasites., Competing Interests: The authors declare that they have no conflict of interest.

Blastocystis is a prevalent eukaryotic microorganism that has been frequently reported from the stool samples of human and animals, all over the world. The prevalence of this parasite has close correlation with the climate and hygiene situation of studied populations. For the current study, English and Persian databases were searched for the following terms: "Blastocystis hominis", "Blastocystis sp.", "Intestinal Parasites", "Prevalence" and "Iran". In order to data analysis and assess the possibility of publication bias, STATA v11 and Begg's Funnel plot were employed respectively. From 605 searched cases, 86 publications met our criteria for the analysis. The pooled prevalence of Blastocystis was 9.1% (95%CI: 8.2%, 10.1%). The results represented that the prevalence of the parasite using molecular techniques or cultivation followed by molecular techniques (20.89%) was higher than only microscopical examination (8.96%). The prevalence rate of Blastocystis according to human development index was 7.6%, 7.2%, 11.9%, 13.4%, and 3.3% for ranks 1 to 5, respectively. This is the first study signifying the prevalence of Blastocystis regarding the human development index and climate conditions. The findings represented that although human development index, low levels of socioeconomic situations together with appropriate climate such as sufficient humidity and high temperature increase the chance of transmission of Blastocystis in a society., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Aim: The aim of the present study was to simultaneously investigate parasitic contamination of treated wastewater and downstream vegetable farms that are irrigated with treated sewage, during a year., Background: (Oo) Cysts and eggs of parasites are resistant to most of routine wastewater treatment process. Irrigation of vegetables farms with either treated wastewater or illegally use of raw wastewaters enhances the risk of contamination with enteric pathogens., Methods: The treated wastewater samples were taken after chlorination from a wastewater treatment plant located at the south of Tehran. In addition, 60 vegetable samples (5 samples from each farm) were collected from the selected downstream farms that routinely used treated wastewater for irrigation of crops. Parasitological tests were performed using Ziehl-Neelsen, conventional lugol's iodine staining and direct microscopical examination., Results: Parasites including free living larvae, eggs of Toxoascaris leonina , egg of Toxocara sp. Trichuris sp, Trichostrongylus sp and amoeboid trophozoite were seen in 5/12 (41.7%) of vegetable samples gathered during a year. There was no statistically significant correlation between the season and parasitic contamination of the vegetables (P= 1). Furthermore, parasitic contamination was observed in 7/12 (53.8%) of treated wastewater samples. The correlation between season and parasitic contamination of treated wastewater was evaluated that the results showed a higher contamination of treated wastewater in spring and autumn (P<0.05). Fisher's exact test also showed that there was no significant correlation between parasitic contaminations of vegetable samples and treated wastewater according to seasonal change., Conclusion: The results showed parasites in both treated wastewater plant and downstream crops farms that suggests the public health importance of the quality of water resources that routinely used for irrigation of vegetable farms., Competing Interests: The authors declare that they have no conflict of interest.

Background: Human-infecting microsporidia are a group of spore-forming eukaryotic microorganisms that can infect both animals and humans. Recent evidences indicate waterborne transmission of microsporidia spores to human via either drinking water or irrigation of vegetable farms with contaminated water resources. The current study aimed to evaluate the presence of human-infecting microsporidia in treated wastewater (TW) and vegetable farms irrigated with treated wastewater during a year., Methods: Totally, twelve samples of each treated wastewater and vegetables were collected. In order to recover microsporidia spores, filtration using cellulose nitrate membrane (pore size 0.4 μm) and sedimentation were employed. DNA extraction was performed for all samples and genus/species were characterized using specific primers. In order to characterize genotypes, ITS fragment of E. bieneusi was amplified, sequenced and compared in GenBank database. Phylogenetic tree was employed to analysis the probable correlation between obtained genotypes with those E. bieneusi genotypes, which were previously obtained from human and animals from same region., Results: After nested PCR, expected fragments of E. bieneusi and Encephalitozoon spp were observed among 5/12 (41.7%) and 1/12 (8.33%) of vegetable samples, respectively. From total of 12 TW samples, expected fragments of E. bieneusi and Encephalitozoon spp were amplified among 7/12 (53.8%) and 1/12 (8.33%) of TW samples, respectively. Genotypes D and E were characterized from both TW and vegetables samples. Phylogenetic analysis showed close-relationship between E. bieneusi from TW and vegetable samples with E. bieneusi from animals and humans obtained from the same region., Conclusion: Our findings suggested the key role of animals in epidemiology of zoonotic transmission of E. bieneusi. Moreover, our findings revealed the occurrence of human-infecting microsporidia in treated wastewater because of either insufficiency of treatment process or distribution of microsporidia spores in wastewater treatment plant via animals., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

The present study tested 80 samples of municipal, geothermal and recreational water samples for the occurrence of waterborne free living amoebae (FLA) including Acanthamoeba, Balamuthia mandrillaris, Vahlkampfiids and Vermamoeba in Semnan province, North half of Iran. Four sets of primers including JDP1,2 primers, ITS1,2 primers (Vahlkampfiids), 16S rRNABal primers (Balamuthia mandrillaris) and NA1,2 primers (Vermamoeba) were used to confirm the morphological identification. From the 80 water samples tested in the present study, 16 (20%) were positive for the outgrowth of free living amoebae based on the morphological page key. Out of the 34 municipal water samples, 7 (20.6%) were positive for outgrowth of Free living amoeba, belonging to Vermamoeba, Naegleria and Acanthamoeba using molecular tools. Three out of the six investigated hot springs were also contaminated with Naegleria spp. Sequencing of the ITS1,2 region of the Vahlkampfiid isolates revealed the highest homology with N. gruberi (2 isolates), N. australiensis (1 isolate) and N. pagei (3 isolates). This is the first report of N. gruberi in the country. Using morphological and molecular analysis, Balamuthia mandrillaris was undetected in all the water samples. The present study further confirmed the occurrence of potentially pathogenic waterborne free living amoebae in habitats with high human activity. It is of utmost importance that more studies are conducted to evaluate the niches of B. mandrillaris and N. fowleri in Iran and worldwide. Such investigations regarding the relevance of FLA as a hazard to humans, should be brought to the notice of the health authorities., (Copyright © 2017. Published by Elsevier Inc.)