Seroprevalence of Toxoplasma gondii in Sheep,Cattle and Horses in Urmia North-West of Iran

Background

Toxoplasma gondii is a zoonotic protozoan parasite found worldwide and responsible for major economic losses in most classes of livestock. This study was aimed to determine the prevalence of T. gondii infection in sheep, cattle and horses in Urmia, north-west of Iran, using MAT.

Methods

Blood samples of 276 livestock and 26 horses were collected from July 2009 till April 2010. The data were analyzed by the Chi-square, Fisher''s Exact and Cochran''s and Mantel-Haenszel Tests. The level of significance was set at P<0.05.

Results

Thirty-three (21.1%) sheep, 2 (1.6%) cattle and 3 (11.5%) horses were seropositive to T. gondii. Analysis showed that sheep were 15 times more likely to be seropositive comparing to cattle also 2 times more likely to be seropositive than horses.

Conclusion

This study showed seroprevalence of equine T. gondii infection with a considerable rate in sheep in Urmia, northwest of Iran. More comprehensive studies on livestock toxoplasmosis are required for further analysis of the parasite reservoir for human infection.     

Seroprevalence of Neospora caninum Infection in Dairy Cattle in Tabriz,Northwest Iran

Background

The aim of this study was to determine the seroprevalence of antibody to Neospora caninum in healthy and aborted dairy cattle in Tabriz, capital of East-Azarbaijan in northwest of Iran.

Methods

In this cross-sectional study serum samples were collected from 266 healthy and aborted Holestein-Feriesisnc cows from September 2008 to August 2009. The sera were analyzed to detect of antibody against N. caninum using the commercially ELISA kit.

Results

Seroprevalence of antibody to N. caninum was 10.5% in Tabriz dairy cattle. Also the abortion rate in all cattle sampled was 33.6% but percentage of seropositive aborted cattle was 18.4%.

Conclusion

Neosporosis could be one of the possible causes of abortion in dairy cattle in Tabriz and regarding the distribution in dogs as definitive host for the parasite, further studies in dog and cattle are recommended.     

Molecular Epidemiology of Cryptosporidiosis in Iranian Children,Tehran, Iran

Background

Cryptosporidium is a worldwide protozoan parasite and one of the most common causes of infection and diarrhea in humans and cattle. The aim of the present study was determination of subtypes of Cryptosporidium among children with diarrhea in Tehran by sequence analysis of the highly polymorphic 60-kDa glycoprotein (GP60) gene.

Methods

Fecal samples were collected from 794 diarrheic children. Initial identification of Cryptosporidium was carried out on stool samples by Ziehl-Neelsen acid-fast staining method. DNA was extracted from positive microscopically samples and Cryptosporidium genotypes and subtypes were determined, accordingly.

Results

Out of 794 collected samples, 19 (2.40%) were positive for Cryptosporidium oocysts. Sequences analysis of GP60 gene showed that 17 (89.47%) of the positive isolates were Cryptosporidium parvum and 2 (10.52%) were C. hominis. All subtypes of C. parvum isolates belonged to allele families IIa (6/17) and IId (11/17). The most common allele in all 17 isolates belonged to IId A20G1a (41.18%). A22G1 (IF) subtype was detected in two C. hominis isolates of the children.

Conclusion

The predominancy of C. parvum species (specially, IId A20G1a subtype) in current study underlines the importance of zoonotic Cryptosporidium transmission in Iran.     

Evaluation of Benzimidazole Resistance in Haemonchus contortus Using Comparative PCR-RFLP Methods

Background

In order to deworm the ruminants especially of sheep in Iran, consumption of benzimidazoles has more than 2 decades history and today farmers are using imidazothiazoles, macrocyclic lactones and mostly benzimidazole compounds (BZs) to treat infected farm animals. It has been demonstrated that the most common molecular mechanism leading to BZsresistance in Haemonchus contortus is a single mutation at amino acid 200 (phenylalanine to tyrosine) of the isotype 1 of beta tubulin gene. According to the report of such mutations in Iranian Teladorsagia circumcincta isolates with Restriction Site Created PCR-RFLP, we decided to evaluate the frequency of such mutations in H. contortus in three different geographical areas of Iran.

Methods

A total of 102 collected adult male H. contortus were evaluated with PCR-RFLP (using PSP1406I as restriction enzyme). By means of a second step to compare function of different methods and to increase sensitivity of detection mechanism, a third of samples were examined by another PCR-RFLP method (using TaaI as restriction enzyme) and finally beta tubulin gene of two samples was sequenced.

Results

All of samples were detected as BZss homozygote. Finally, beta tubulin gene sequencing of two samples showed no point mutation at codon 200.

Conclusion

It seems that BZresistance of H. contortus in Iran is not a serious problem as anticipated before.     

Identification and Genetic Variation of Fasciola Species from Tabriz,North- Western Iran

Background

Fascioliasis is considered as the most important helminthic infection of cattle and sheep. Traditional approaches using morphological and biologic characters cannot cause a certainty in the accurate and precise identification and intra-specific differences of Fasciola spp. In this study, we identified Fasciola species using ITS-1 marker and described genetic variation of each species of the parasite in isolates from Tabriz slaughterhouse in West Azerbaijan Province, north- western Iran.

Methods

Overall, 100 samples (50 from sheep and 50 from cattle) morphologically detected as Fasciola worms were studied for identification of Fasciola species by PCR-RFLP method and intra-species variation of the parasite using RAPD-PCR technique.

Results

A region of approximately 460bp in all samples was successfully amplified. There were no identifiable variations among the size of PCR products. Two and three fragments in samples correspond to F. hepatica and F. gigantica was seen, respectively, through PCR-RFLP method. No difference was seen in digestion pattern according to host (sheep or cattle). Different types of each species of the parasite was observed using RAPD-PCR technique.

Conclusion

We could have an estimate of frequency of F. hepatica and F. gigantic and different genotypes of the parasite in isolates from one locality in north- western of Iran. By extension of such studies in future to other animal hosts (buffalo and goat) and including more regions to sampling, the reliability of the results and their application for control programs in zoonotic diseases will be increased.     

Absence of Asymptomatic Malaria Infection in Endemic Area of Bashagard District,Hormozgan Province,Iran

Background

A successful malaria elimination program calls for enough attention to parasite carriers, especially asymptomatic malaria, as well as the diagnosis and treatment of clinical cases. Asymptomatic malaria is an infection that patients do not show any symptom; thus, these patients play critical role in the concept of an elimination program. The current investigation was conducted to evaluate the presence of these cases in Bashagard District, formerly a high malaria transmission area in Hormozgan Province, Iran.

Methods

Blood samples (n = 500) were collected from symptomless individuals residing in Bashagard to evaluate Plasmodium infection by using microscopic, serological and nested-PCR techniques.

Results

Regarding the microscopic and nested-PCR analysis, no asymptomatic infection was detected among studied individuals. Totally, 1% of the studied population (5 of 500) had anti PvMSP-1₁₉-specific IgG antibody; however, only 0.2% (1 of 500) of the individuals was seropositive to recombinant PfMSP-1₁₉, using ELISA.

Conclusion

This study showed no asymptomatic malaria infection in the studied population; hence malaria elimination is feasible and can be successfully carried out in this region.     

Seroprevalence of Acanthamoeba Antibodies in Rheumatoid Arthritis Patients by IFAT,Tehran, Iran 2007

Background

This preliminary study was conducted to discriminate the prevalence of Acanthamoeba antibodies in rheumatoid arthritis (RA) patients and healthy controls to analyze the correlation between these two groups.

Methods

From October 2006 to August 2007 a total of 121 serum samples from RA patients attending the Rheumatolgy Department at Shariati Hospital in Tehran were obtained and stored at -20°C until using by indirect fluorescent-antibody test (IFAT). RA was diagnosed according to the American Collage of Rheumatology classification criteria. The organism used in this study was isolated from various water resources in Tehran, Iran cultured axenically and then went on a PCR assay based on 18S rRNA to identify the genus Acanthomoeba. Indirect immunofluorescence antibody (IFA) staining of serum samples was carried out to detect anti Acanthomoeba antibodies.

Results

In culture, out of 22 samples, 13(59%) were grown in xenic but only two in axenic medium. PCR amplified a 904bp fragment, specific for Acanthamoeba. Of examined serum samples, Acanthamoeba antibodies were present in 70 (57.8%) and 52 (41.2%), respectively. The highest titer of antibodies (1:320) was detected in one patient with RA.

Conclusion

Our study supports the hypothesis that some parasitic microorganisms can involve and contribute toward the development of rheumatoid syndromes.     

Molecular and Seroepidemiological Survey of Visceral Leishmaniasis among Humans and Domestic Dogs in Mazandaran Province,North of Iran

Background

New cases of visceral leishmaniasis (VL) have been reported recently in some parts of Mazandaran Province, north of Iran where the first human case of VL was reported in 1949. This study aimed to determine the present status of Leishmania infantum infection among humans and domestic dogs using serological and molecular methods in central parts of Mazandaran Province.

Methods

In this cross-sectional study, blood samples were randomly collected from 402 humans and forty-nine domestic dogs throughout 2009 and 2010 in the central part of Mazandaran Province including Semeskadeh and Kiakola districts where recent cases of human visceral leishmaniasis had been reported there. All the collected samples were tested by direct agglutination test (DAT) for the detection of anti-Leishmania infantum antibodies as well as convenience PCR assay on whole blood samples for detection of leishmanial infection and identification of Leishmania species.

Results

None of 402 collected human (402) and dog (49) blood samples showed anti Leishmania infantum antibodies at titers 1:3200 and 1:320 as cut-off values of DAT, respectively but only 2 of domestic dogs (4.1%) were found PCR-positive corresponding to L.infantum.

Conclusion

This study confirms the circulation of L. infantum at least among domestic dogs and highlights the sporadic pattern of VL in the studied areas. Further investigations regarding to sand flies fauna and wild canines as reservoir hosts of the disease, are recommended.     

First Detection of Nosema ceranae,a Microsporidian Protozoa of European Honeybees (Apis mellifera) In Iran

Background

Nosemosis of European honey bee (Apis mellifera) is present in bee colonies worldwide. Until recently, Nosema apis had been regarded as the causative agent of the disease, that causes heavy economic losses in apicultures. Nosema ceranae is an emerging microsporidian parasite of European honeybees, A. mellifera, but its distribution is not well known. Previously, nosemosis in honeybees in Iran was attributed exclusively to N. apis.

Methods

Six Nosema positive samples (determined from light microscopy of spores) of adult worker bees from one province of Iran (Savadkouh- Mazandaran, northern Iran) were tested to determine Nosema species using previously- developed PCR primers of the 16 S rRNA gene. As it is difficult to distinguish N. ceranae and N. apis morphologically, a PCR assay based on 16 S ribosomal RNA has been used to differentiate N. apis and N. ceranae.

Results

Only N. ceranae was found in all samples, indicating that this species present in Iran apiaries.

Conclusion

This is the first report of N. ceranae in colonies of A. mellifera in Iran. It seems that intensive surveys are needed to determine the distribution and prevalence of N. ceranae in different regions of Iran.     

Heavy Worm Burden of Moniliformis moniliformis in Urban Rats with Histopathological Description

Background

Due to scarcity of human reports, we took advantage of the heaviest infection of M. moniliformis in rats, to describe histopathological and microanatomical valuable useful keys while confronting human occurrences.

Methods

Samples were obtained from captured rats in Tehran, capital of Iran, during two decades. Tissue sections were performed through hematoxylin and eosin staining to describe histopathological changes in rat''s intestines.

Results

Totally, nine rats were found infected with M. moniliformis amongst 272 obtained rats. Heavy infection has been distinguished in 2 individuals with parasite burden of 141and 73 adult worms. Cross sections of worms within the lumen show mucosal thickness, infiltration of eosinophilic leukocyte and increase in goblet cells.

Conclusion

Beyond the uncommonness of human infection with M. moniliformis unitended infections should not be ignored. Abundance of rats and roaches as definite and intermediate hosts must be considered particularly in countries with poor hygiene.     

Establishment of Hymenolepis diminuta Life Cycle to Provide Parasite Mass Production

Background

The main object of this experimental work was to practise laboratory production both adult and the larval stage of Hymenolepis diminuta with conventional modification to make further studies easier.

Materials & Method

Adults H. diminuta were collected from urban rats in Tehran, Iran. The beetles became infected using blended gravid segments with flour as bait. Cysticercoids have been saved after precise dissection of invertebrate hosts. The exposure of infected beetles to laboratory rats was performed to establish the life cycle.

Result

Out of 57 collected rats, three rats were infected with H. diminuta. Almost all exposed beetles found infected with the larval stage of parasite. About one-month later H. diminuta eggs were seen in stool examination of laboratory rats.

Conclusion

Rare human occurrence of H. diminuta along with light level of clinical manifestation of this parasite, underestimate the concerns toward its public health importance. Nowadays, various field of studies, such as biochemistry with special focuses on the capability of H. diminuta tegument absorption have performed apart from parasitological views alone. In the present study, establishment of this parasite life cycle has practically provided the access of adult and cysticercoid stages of the tapeworm in further researches.     

Molecular and Biomorphometrical Identification of Ovine Babesiosis in Iran

Background

Ovine babesiosis is the most important haemoparasitic tick-borne disease of small ruminants in Iran caused by Babesia ovis, B. motasi, and B. crassa. The aim of this study was to characterize the species of ovine Babesia species isolated from different geographical region of Iran.

Methods

One hundred fifty four blood samples collected from animals, which demonstrated the pale mucous membranes or hyperthermia. The specimens were transferred to the laboratory and the blood smears stained with Geimsa, the morphological and biometrical data of parasite in any infected erythrocyte have been considered. Extracted DNA from each blood samples were used in PCR and semi nested- PCR in order to confirm the presence of the species.

Results

Microscopical observation on 154 blood smears determined 38 (24.67%) and 40 (26%) samples were infected by Babesia and Theileria respectively. The mixed infections occurred in four (2.6%) samples. The results of the PCR assays showed nine (5.85%), 81 (53%) and 18 (11.7%) were distinguished as Babesia, Theileria and mixed infection, respectively. Semi nested- PCR did not confirm the presence of B. motasi.

Conclusion

The causative organism of many cases of haemoprotozoal diseases, which recorded in previous studies, could be B. ovis or Theileria lestoquardi. The result confirmed that B. ovis was only species which causes babesiosis in the study areas. It seems that the biometrical polymorphisms could exist in B. ovis in Iran. This polymorphism could be a main problem in differentiation between B. ovis and B. motasi and it could be dissolved by specific PCR analysis.     

Toxoplasma Infection in Schizophrenia Patients: A Comparative Study with Control Group

Background

Schizophrenia is a serious, chronic, and often debilitating neuropsychiatric disorder. Its causes are still poorly understood. Besides genetic and non-genetic (environmental) factors are thought to be important as the cause of the structural and functional deficits that characterize schizophrenia. This study aimed to compare Toxoplasma gondii infection between schizophrenia patients and non-schizophrenia individuals as control group.

Methods

A case-control study was designed in Tehran, Iran during 2009-2010. Sixty-two patients with schizophrenia and 62 non-schizophrenia volunteers were selected. To ascertain a possible relationship between T. gondii infection and schizophrenia, anti-Toxoplasma IgG antibodies were detected by indirect-ELISA. Data were statistically analyzed by chi- square at a confidence level of 99%.

Results

The sero-positivity rate among patients with schizophrenia (67.7%) was significantly higher than control group (37.1) (P <0. 01).

Conclusion

A significant correlation between Toxoplasma infection and schizophrenia might be expected.     

Occurrence of Thermotolerant Hartmannella vermiformis and Naegleria Spp. in Hot Springs of Ardebil Province,Northwest Iran

Background

Geothermal waters could be suitable niches for thermophilic free living amoebae including Naegleria and Hartmannella. Ardebil Province, northwest Iran is popular for having many hot springs for recreational and health purposes activity. The present research is the first molecular based investigation regarding the presence of Naegleria and Hartmannella in the hot springs of Ardebil Province in Iran.

Methods

Overall, 30 water samples were taken from waters of thermal hot springs in Ardebil Province, Iran during 2010-2011. All collected samples were transferred to Dept. of Parasitology and Mycology, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Cultivation of concentrated water samples was performed using culture-enrichment method. Cloning of the target amoebae was obtained and morphological and molecular analysis was done using page key combined with two sets of primers, respectively. Sequence analysis and homology search was used for strains identification.

Results

Of 30 water samples, 8 (26.7%) were positive for thermotolerant Vahlkampfiids and Hartmannella based on morphological characteristics of vegetative form and double walled cysts. Cloning of the target amoebae were done successfully. Sequencing of the positive isolates revealed that the strains belonged to Naegleria (N. carteri and N. spp) and H. vermiformis.

Conclusion

The result highlights a need for improved filtration and disinfection and periodic monitoring of recreational thermal waters in order to prevent disease related to free- living amoebae. This is the first comprehensive molecular study of thermophilic Naegleria and Hartmannella in hot springs of Iran.     

Isolation of Free-Living Amoebae from Sarein Hot Springs in Ardebil Province,Iran

Background

Free-living amoebae (FLA) are a group of ubiquitous protozoan, which are distributed in the natural and artificial environment sources. The main aim of the current study was to identify the presence of FLA in the recreational hot springs of Sarein in Ardebil Province of Iran.

Methods

Seven recreational hot springs were selected in Sarein City and 28 water samples (four from each hot spring) were collected using 500 ml sterile plastic bottles during three month. Filtration of water samples was performed, and culture was done in non-nutrient agar medium enriched with Escherichia coli. Identification of the FLA was based on morphological criteria of cysts and trophozoites. Genotype identification of Acanthamoeba positive samples were also performed using sequencing based method.

Results

Overall, 12 out of 28 (42.9%) samples were positive for FLA which Acanthamoeba and Vahlkampfiid amoebae were found in one (3.6%) and 11 (39.3%) samples, respectively. Sequence analysis of the single isolate of Acanthamoeba revealed potentially pathogenic T₄ genotype corresponding to A. castellanii.

Conclusion

Contamination of hot springs to FLA, such as Acanthamoeba T₄ genotype (A. castellanii) and Vahlkampfiid amoebae, could present a sanitary risk for high risk people, and health authorities must be aware of FLA presence.     

Investigation of Possible Correlation between Giardia duodenalis Genotypes and Clinical Symptoms in Southwest of Iran

Background

Giardia duodenalis is one of the most important human enteric parasites throughout the world. Clinical symptoms of this parasite vary from asymptomatic infection to chronic diarrhea. Still it is not clear, whether different types of pathogenesis are due to different strains of organism or to variable host factors. The purpose of this study was to investigate possible correlation of clinical symptoms with assemblages among symptomatic and asymptomatic cases collected from southwest of Iran.

Methods

Fecal samples were collected from 100 symptomatic and asymptomatic cases, which were positive for G. duodenalis. The samples were subjected to semi-nested PCR and RFLP for gdh gene.

Results

Among symptomatic patients, 54% had mixed genotypes AII and BIII, 28% and 18% of samples indicated assemblages BIII and AII, respectively. In contrast, among asymptomatic cases, 64%, 26% and 10%samples had mixed genotypes, BIII and AII assemblages, respectively. Statistical analysis using Chi- Square test showed that there was no significant correlation between assemblage and clinical symptoms in current study.

Conclusion

High prevalence of mixed infection in both groups may affect this conclusion, therefore further study in more details are necessary to clarify these finding. Additionally, it is important to carry out investigations regarding human host factors as well.     

Development of Sensitive Detection of Cryptosporidium and Giardia from Surface Water in Iran

Background

The protozoan parasites Cryptosporidium spp. and Giardia are known to occur widely in both raw and drinking waters. They are two of the causative agents of waterborne outbreaks of gastroenteritis throughout the world. In the present study, a PCR assay and FA were developed for detection of Cryptosporidium oocysts and Giardia cyst in environmental samples.

Methods

We have detected Cryptosporidium spp. oocysts and Giardia cysts in seeded and unseeded environmental water samples by PCR method. Water samples were spiked with oocysts (50, 100,300,500) and filtrated with a 1.2-µm pore size cellulose nitrate and follow by DNA extraction and purification by QIAamp DNA mini kit. Nested-PCR assay amplified an 850 bp fragment of 18s rRNA gene specific for Cryptosporidium and 435 bp fragment of glutamate dehydrogenase (GDH) target gene for Giardia. Also many river water from north of Iran, be checked by these methods.

Results

Cryptosporidium and Giardia DNAs were detected in seeded water sample and Giardia was detected in all 5 water samples from river in north of Iran by nested- PCR and FA. Also in one river water sample, Cryptosporidium was detected.

Conclusion

This protocol is effective for detection of these waterborne parasites in treated and untreated water samples. This study can also serve as a platform for further investigations and research water source in Iran.     

In vivo Susceptibility of Plasmodium vivax to Chloroquine in Southeastern Iran

Background

Plasmodium vivax is the predominant species causes of malaria with about 90% total annual reported malaria in Iran. This study conducted to determine the susceptibility of Plasmodium vivax isolates to chloroquine in Sistan and Balochistan Province, southeastern Iran.

Methods

A total 270 subjects with symptomatic malaria and confirmed P. vivax infection completed the designed 28-day in vivo study. The thick and thin film blood smears were screened for malaria parasites by microscopy. The nested PCR was applied using the Plasmodium 18 subunit ribosomal ribonucleic (Ssr RNA) genes for detecting mixed infections and diagnosis of parasites in the samples with low parasite on days 0, 5, 6, 7, and 28.

Results

P. vivax was cleared in 15%, 50%, 95%, and 100% of patients on days 1, 2, 3, 4 respectively by microscopy assessment. Six patients were exhibited specific P. vivax band in nested PCR on day 5. No recurrence was observed on days 7, 14 and 28. Mean (±standard deviation) parasite clearance time was 2.41 (±0.8) days.

Conclusion

P. vivax is still susceptible to chloroquine in Southeatern Iran. This finding is compatible with results of neighboring countries Pakistan and Afghanistan.     

Species Identification of Birds Nasal Trichobilharzia in Sari,North of Iran

Background

Cercarial dermatitis is known as an endemic parasitic disease in North of Iran, a hypersensitive skin reaction to the penetration of nonhuman schistosome larvae into human skin. In recent studies in this region, final and intermediate hosts were recognized and Trichobilharzia was identified as the main causative agent of cercarial dermatitis in this region, but to date the parasite species haven''t been identified. Therefore this study was performed to species identification of nasal Trichobilharzia in infected birds for the first time.

Methods

A total of 45 Anas clypeata birds identified as final host, were collected from Sari in North of Iran and infected nasal tissues analyzed using molecular techniques. Genomic DNA was isolated by phenol/chloroform extraction method and ITS region of rDNA were amplified with specific primers its5Trem and its4Trem, then sequenced area were compared with existing records in GenBank.

Results

Twelve samples were infected with Trichobilharzia and results of PCR reaction indicated that all of them belonged to T. regenti. The sequence alignment of present work isolates and those deposited in GenBank showed differences in nucleotide sequences of repeat region in ITS1.

Conclusion

Trichobilharzia regenti is the most frequent parasite of Anatid birds in North of Iran. This corresponds to the distribution of this parasite along the flyway of migratory birds, which annually migrate from Siberia and northern countries of Caspian Sea to wintering areas in southern regions of it.     

Seroprevalence of Toxoplasma gondii in Pregnant Women and Bioassay of IgM Positive Cases in Zanjan,Northwest of Iran

Background

The aim of this study was to determine the Toxoplasma antibodies in pregnant women in Zanjan, by ELISA method.

Methods

Blood samples were taken from 500 pregnant women referred to the health centers of Zanjan City, North West Iran, IgM and IgG titers were primarily evaluated. The collected data were analyzed with SPSS 11.5 using Chi-Square test.

Results

Anti Toxoplasma IgM and IgG were positive in 1.4% and 37.2% respectively. Seropositive subjects were more frequently seen in those with >30 years old compared to younger women (<20 years old). No significant relationship was found between the seroprevalence of T. gondii infection and level of education, residence area, history of abortion and gestational age.

Conclusion

The rate of IgM positive was low; however, a large number of the studied population were IgG positive, indicative of having a latent infection due to the past exposure to Toxoplasma parasite in this region.