Genotyping of hydatid cyst isolated from human and domestic animals in ilam province, western Iran Using PCR-RFLP

Hydatidosis or cystic hydatid disease is one of the most important diseases in human and animals. Identification of strains is important for improvement of control and prevention of disease. The aim of this study was to determine the strains isolated from human and domestic animals in Ilam Province, Iran, using PCR-RFLP method. Respectively, 30 and 4 animal and human hydatid cysts were collected from different slaughterhouses and hospitals of the province. Protoscolices were separated and their DNA genome was extracted by extraction kit. rDNA-ITS1 of each isolated samples was duplicated by BD1 [Forward] and 4s [Reverse] Primers. PCR products were studied by electrophoresis and then were digested using TaqI, HpaII, Rsal and Alul restriction enzymes. RFLP products were studied using electrophoresis on 1% agar gel. A fragment of l000bp was produced from amplification of rDNA-ITS1 of protoscolices using PCR method. After digestion of PCR product by Alul enzyme, 200bp and 800bp, by Rsal, 655bp and 345bp and by HpaII 700bp and 300bp sizes were obtained. TaqI enzyme had no change in fragment size and it remained l000bp. Considering the method. Ham strains was specified as E. granulosus sensu stricto [G1-G3]. Although sheep strain [G1] is dominated in human and different animal in Iran and the world, but more efforts should be done to clarify the true genotype of Ilam strains specified as E. granulosus sensu stricto [G1-G3]

Assessing the effect of highly cited papers on the impact factor of journals in the field of public health

The aim of this study was to appraisal the effect of highly cited papers in the field of public health and find out whether the unusual citations affect the ranking order of the journals in this field or not. A total number of 142 journals titles were listed in Journal Citation Report [ISI Thomson] in the field of "Public, Environmental and Occupational Health". All but one of them had published papers at least for a year from 2009 to 2010. Journal title, number of citations and publication year of 45685 papers were collected from ISI web of knowledge database at December 25, 2011. About half of the papers [23226] had no citations and 89.4% [40835] had less than 6 citations. We concluded that the ranking of journals in the field of public health is not affected by the individual papers with unusual number of citations

Seroprevalence of human hydatidosis using ELISA method in Qom Province, Central Iran

The objective of this study was to determine the prevalence of cystic echinococcosis [CE] in Qom Province, central Iran using ELISA test. Overall, 1564 serum samples [800 males and 764 females] were collected from selected subjects by randomized cluster sampling in 2011-2012. Sera were analyzed by ELISA test using AgB. Before sampling, a questionnaire was filled out for each case. Data were analyzed using Chi-square test and multivariate logistic regression for risk factors analysis. Seropositivity was 1.6% [25 cases]. Males [2.2%] showed significantly more positivity than females [0.9%] [P= 0.03]. There was no significant association between CE seropositivity and age group, occupation, and region. Age group of 30-60 years encompassed the highest rate of positivity. The seropositivity of CE was 2.1% and 1.2% for urban and rural cases respectively. Binary logistic regression showed that males were 2.5 times at higher risk for infection than females. Although seroprevalence of CE is relatively low in Qom Province, yet due to the importance of the disease, all preventive measures should be taken into consideration

[Prevalence and severity of animal Fasciolosis in six provinces of Iran]

Fasciolosis is one of the most important parasitic disease common among both humans and livestock. Considering the health and economic importance of the disease, an understanding of the epidemiology of Fasciolosis is highly crucial. This study aimed to evaluate the prevalence and severity of Fasciola infection in animals from different geographical regions of Iran during 2009-10. In this descriptive study, 11100 livers taken from slaughtered sheep and cattle were carefully examined for Fasciola parasites at the six industrial slaughterhouses of East Azerbaijan, Khorasan-Razavi, Khuzestan, Fars, Mazandaran and Markazi provinces. All Fasciola parasites isolated from the livers of infected animals were transferred to the laboratory, and then the parasite species were identified and counted. Finally, the frequency distribution and the severity of infection were analyzed. In this study, 1.10% of the total sheep and cattle slaughtered in six industrial slaughterhouses were found positive for Fasciolosis. The severity of Fasciola in sheep and cattle livers was 7.77 +/- 0.42 and 15.24 +/- 1.78, respectively. Khorasan Razavi and Fars provinces had the highest [14.54 +/- 3.16] and lowest [7.75 +/- 0.79] severity of infection, respectively. Rresults of the study show a reduction in the prevalence and severity of Fasciolosis in sheep and cattle. But considering the importance of the disease and its endemicity, the preventive measures should be taken against the animal and human Fasciolosis in Iran

prevalence of honorary and ghost authorships in Iranian biomedical journals and its associated factors

The purpose of this study was to detect the prevalence of ghost and honorary authors and its determinant factors in bio-medical journals of Iran. The study was done in 2009-10 in Tehran, Kerman, and Iran Medical Universities, Iran. We contacted the first or corresponding authors of the papers had published papers in the recent two issues of Iranian Journal of Public Health, Journal of Kerman University of Medical Sciences, and Tehran University Medical Journal. They explored the role of each coauthor and others who had done mouthing for the paper. Then, according to ICMJE criteria, we counted how many of them are real, honorary or ghost author. For the analysis, we utilized two databases. One included articles as the records and the other included authors as the records. From 124 articles, with 536 authors, 301 [56.1%] were honorary authors. Each article had 4.35 authors on average, while 2.4 of them were honorary authors. The percentage of honorary author in basic science articles was about 6% more than the articles of clinical sciences. Moreover, 89% of articles had at least one honorary author. About 20% of all articles had more than three honorary authors. Besides, 25 [21.43%] authors confessed they had colleague[s] omitted from the authors list, while only one [0.81%] of them met the authorship criteria. The percentage of agreement between the corresponding and the remaining authors on the number of honorary of the authors was about 47.4% [Kappa= 0.27, P= 0.01]. It seems that the present data might assist the authorities to make a decisive decision on amending the process of authorship in Iran

Molecular diagnosis of Strongyloides stercoralis infection by PCR detection of specific DNA in human stool samples

Strongyloidiasis is mostly an asymptomatic infection and diagnosis of latent infections is difficult due to limitations of current parasitological and serological methods. This study was conducted to set up a PCR-based method for molecular diagnosis of Strongyloides stercoralis infection by detection of copro-DNA in stool samples. A total of 782 fresh stool samples were collected and examined by agar plate culture. Among those sixteen stool samples, which confirmed to be infected with S. stercoralis were examined as positive control to set up each single and nested PCR, using two primer sets designing to amplify partial ribosomal DNA of S. stercoralis genome. Since, single PCR method yielded higher efficacy in detecting positive samples, in the second step, 30 stool samples, which found negative for S. stercoralis by agar plate culture of single stool sample, were examined by single PCR. Data analysis was performed using McNemar's chi[2] test, with consideration of a P-value of <0.05 as indication of significant difference. In amplification of DNA extracted from stool samples, single PCR detected S. stercoralis DNA target in all 16 positive samples, while nested PCR amplified DNA in only 75% of samples. In the second step, single PCR amplified S. stercoralis extracted DNA in 5 out of 30 samples which were negative by coproculture. Single PCR method amplifying a short [100bp] target represented more efficacies for detection of S. stercoralis in faecal examination compared to agar plate culture and nested PCR, which amplified longer target

Establishment of Hymenolepis diminuta life cycle to provide parasite mass production

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. 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. 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. 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

Evaluation of the stability of coated plates with antigen at different temperatures and times by ELISA test to diagnose fasciolosis

Considering that ELISA method presently is the test of choice for diagnosis of fasciolosis, the present study was undertaken to evaluate the maximum validity of coated plates at different temperatures and different times during one year of evaluation. Serum samples of patients infected with fasciolosis [n=10], hydatidosis [n=5], toxocariasis [n=5], and negative control sera [n=5] were examined. Two series of plates were considered. The first series were coated with Fasciola homogenate Ag 12 micro g/ml, and after some steps were blocked with gelatin and preserved at different temperatures as -80 °C, -20 °C, -4 °C and +4°C. The 2[nd] series were treated under the same criteria but were not blocked with gelatin. Each series were examined by ELISA test from 1[st] month to 12[th] month. Sera with 1:125 dilution, and peroxidase-conjugated goat anti-human IgG diluted 1:10000 were considered optimum. To ease reporting the results and due to many similarities only results related to 1[st], 6[th] and 12[th] months were analyzed and sensitivity, specificity plus cut-off were determined for each series separately. Preserving the coated plates, while unblocked at -80°C for 6-8 months is pertinent and functional and in that case, we can be sure the best out put would be applicable

Molecular characterization of Fasciola hepatica isolates by RAPD-PCR and ribosomal ITS1 sequencing

Understanding genetic structure and status of genetic variation of the Fasciola hepatica populations has important implications for epidemiology and effective control of fasciolosis. The aim of the present study was to genetically characterize F. hepatica isolates from different hosts, using sequence analysis of ribosomal ITS1 and RAPD-PCR. Fifty three adult F. hepaticas were isolated from naturally infected cattle, sheep, buffalo and goat from two regions in Iran. Genomic DNA was extracted from 70% ethanol preserved flukes. RAPD-PCR with a set of arbitrary primers [UBC90 and R151] was used to estimate genetic variation within the species. Ribosomal ITS1 region of the isolates was amplified, using primers specifically designed for this study. Ten samples [4 sheep, 2 cattle, 3 buffaloes and one goat isolate] were sequenced at ITS1 and analyzed, using DNASIS and ClustalW softwares. F. hepatica ITS1 region was amplified successfully for all samples and a band of 470 bp was shown in all cases. Different isolates did not show any significant genetic variations in rDNA-ITS1 as all the sequences showed to be 100% identical. RAPD results of 52 samples, in particular those with UBC90, showed different patterns within F. hepatica isolates of each host. RAPD data for this primer showed three different patterns for each of sheep and cattle isolates and two patterns in buffalo isolates. All the 14 cattle isolates come up with an identical pattern, using primer R151. The study showed the variability of F. hepatica isolates in Iran, using RAPD markers. No intraspecies variation was seen in the Iranian F. hepatica isolates at ITS1 rRNA gene, indicating highly conserved nature of this region

Serological evaluation of EgAgB16 kDa, a recombinant antigen from echinococcus granulosus for diagnosis of human hydatidosis

Regarding that accurate diagnosis of human hydatidosis still needs more investigations, the present study was conducted to clone, express, and evaluate the gene encoding AgB subunits [EgAgB16 kDa] from Echinococcus granulosus [Iranian G1 strain] and its evaluation by ELISA test. DNA was extracted from protoscoleces and was utilized by PCR for strain identification. Total RNA was prepared with RNeasy protect mini kit from E. granulosus [Iranian G1 strain] protoscoleces collected from naturally infected sheep with hydatid cyst. Recombinant AgB16 kDa was produced using pETDuet as vector and evaluated by ELISA method. A panel of sera including hydatid cyst-infected individuals [n=72], healthy individual [n=48], toxoplasmosis [n=4], strongyloidosis [n=4], kala-azar [n=5] and tuberculosis [n=5] were examined using this recombinant antigen. Recombinant protein was purified by affinity chromatography using His-Tag column. After purification, recombinant protein was confirmed by western blot analysis using His Tag monoclonal anti body or hydatid positive human serum. The sensitivity, specificity; positive and negative predictive values were calculated as 93.5%, 95.6%, 96% and 92.9%, in that order. The cut-off point was detected 0.3 for rAgB16. While the produced recombinant AgB16 kDa showed promising results in diagnosing human hydatidosis, but more investigations should be implemented to reach an accurate gold standard

Ocular dirofilariasis, a case report

Accidental infection with animal filarial worms in humans is a dilemma for clinicians and parasitologists throughout the world. To date a variety of such rare parasitoses have been reported mostly in tropics and subtropics. Human dirofilariasis is among those unusual zoonotic infections that occasionally have been observed in the eye and in subcutaneous areas exhibiting with nodule for mation. Filarial worms are transmitted to humans through invertebrate biological vectors such as certain species of mosquitoes. The present report describes a peculiar case of ocular dirofilariasis in a 49-year-old man resident in Iran

Echinococcosis / hydatidosis in Iran

Cystic echinococcosis caused by Echinococcus spp. is considered endemic in Iran. To clarify the present status of hydatidosis in Iran the present review article is presented. Authentic databases and search engines from 1996 onwards were utilized to enquire the situation of the disease in Iran. Human hydatidosis is responsible for approximately 1% of admission to surgical wards and the rate of human infection is 0.6-1.2/100000. The usual order of involvement, i.e. liver, lung, and other organs, respectively is documented here as well. Risk factors include contact with dog, eating vegetable, geophagy and contact with sheep. Dogs play a critical role in transition the hydatidosis. The rate of infection with E. granulosus in stray dogs shows a prevalence of 5% to 49% in different parts of Iran. Followed by sheep with 88% fertilized cysts, camel with 70%, and cattle with 19% have been considered as the most important and the weakest intermediate host of E. granulosus, respectively. Molecular analyses clearly indicate that the camel/dog strain [G6 genotype] of E. granulosus as well as the cosmopolitan, common sheep strain [G1 genotype] occurs in Iran. A wide variety of livestock including sheep, cattle, goat, camel and buffalo also harbor the disease. E. multilocularis another agent of human hydatidosis [alveolar cyst] is reported here as well and from 1946 to 1993, 37 cases of human alveolar echinococcosis were reported from northwestern Iran. Hydatidosis must be considered as a dilemma in Iran because of its endemicity in the country

Survey of urinary schistosomiasis in Iran

To determine the present status of urinary schistosomiasis in Khuzestan Province, southwestern Iran. Urine samples were collected from 3400 villagers residing in the high risk areas in Khuzestan Province, mainly from school children [80%] from 2005-2007. During a sequential visit by our team urine specimens were collected between 10:00am and 14:00pm. Each person was given a prenumbered bottle in the field, and the name of the person including age and sex was entered against the appropriate number on a form kept by the investigating team. In this province, the transmission of Schistosoma haematobium is being successfully interrupted as none of the samples were found positive. Total elimination of urinary schistosomiasis appears to be possible if the health authorities in neighboring areas can be persuaded to adopt a similar strategy of integrated control. The plan for the future is to continue monitoring transmission, by passive surveys in local health centers and active case-finding among schoolchildren, and to continue snail sampling and focal mollusciciding

new primer pair in ITS1 region for molecular studies on echinococcus granulosus

Echinocuccus granulosus, the causative agent of cystic echinococcosis has long been recognized as having a high degree of genetic divergence. The strains characterization seems to be essential for the establishment of a preventive and control strategy in every endemic area. Using DNA based methods for strain /genotype characterizations of E. granulosus have some difficulties, especially access to an efficient and pure concentration of DNA and proper primers. Using grinder method, a pure and high concentration DNA was extracted from 10 human hydatid cysts collected from Isfahan [central Iran] hospitals, and processed for PCR reaction. Using DNASIS, the primers were designed in internal transcribed spacer 1 [ITS1] region, following analysis of 30 E. granulosus nucleotide sequences, extracted from gene bank. This new and specific E. granulosus primer which amplified DNA thoroughly can be applied for molecular studies on echinococcosis

Seroepidemiology of human hydatidosis in Golestan province, Iran

Hydatidosis is one of the most prevalent zoonotic diseases worldwide. So far no survey was conducted to determine the rate of human hydatidosis in Golestan Province, so using IFA and ELISA tests the prevalence of this disease was detected in patients referred to health centers in this province. Totally 1024 serum samples were collected from patients referred to different health centers in 4 cities of Gloestan Province including Gorgan, Gonbad kawoos, Aliabad Katool and Kordkoy. All the sera were examined using IFA and ELISA tests. Twenty four cases [2.34%] were positive for hydatidosis in Golestan Province using IFA, whereas 22 cases [2.15%] showed positivity using ELISA. Gorgan, Gonbadkaoos, Aliabad Katool and Kordkoy demonstrated the rate of positivity as 1.41%, 2.40%, 5.36% and 2.30%, respectively, but no significant difference was seen. As to positivity, there was no significant difference between age groups, sex, different cities and rural or urban life, but a significant different was seen according to job and literacy [P< 0.001]. According to Job and literacy, housewives and illiterates had the highest rate of infection as 3.67% and 3.72%, respectively. As regards residency, urban life showed no significant difference with rural life [2.47% vs. 2.45%]. Age group of 40-49 years old had the highest rate of positivity [3.95%]. Females were more infected than males [3.16% vs. 1.93%]. The rate of prevalence in this province shows somehow a resemblance with the other cities in Iran. Considering the lifestyle in this province a complementary study is suggested in all related cities

Evaluation of Dot-ELISA method using excretory-secretary antigens of fasciola hepatica in laboratory diagnosis of human fasciolosis

Fasciolosis diagnosis, due to low sensitivity of coprological diagnostic method has been challenging for a long period. In this study, Dot-ELISA, one of the simplest and the most sensitive tests in this regard, was evaluated using excretory-secretory antigens of Fasciola hepatica to diagnose human fasciolosis Three groups consisting of patients infected with fasciolosis [n= 95], patients with other parasitic diseases [n= 37] and healthy individuals [n= 40], were implicated in the test. All collected sera were tested by Dot-ELISA using excretory-secretory antigens. Optimal criteria were detected as 1.5 micro g of antigen per dot, serum dilution of 1:320, and anti human IgG conjugate dilution of 1:500. The sensitivity, specificity, positive and negative predictive values were 96.8%, 96.1%, 96.8% and 96.1%, respectively. In conclusion, Dot-ELISA using excretory-secretory antigens could be regarded as a cheap, rapid, antigen and serum conservative diagnostic method in diagnosing fasciolosis

Comparative evaluation of fast enzyme linked immunosorbent assay [fast-ELISA] and standard-ELISA for the diagnosis of human hydatidosis

Fast enzyme linked immunosorbent assay [Fast-ELISA] was compared with the standard ELISA for the diagnosis of human hydatidosis. Seventy serum samples including 30 from hydatidosis patients [surgically confirmed], healthy control individuals not infected with any parasitic diseases [n=/20] and from others with different parasitic infections including, toxocariosis [n=5], fasciolosis [n=5], trichostrongylosis [n=5], and strongyloidosis [n=5] were analysed for anti-hydatid IgG antibodies using sheep hydatid cyst fluid antigen. The sensitivity, specificity, positive and negative predictive values, as well as validity of the test were found as 96.7%, 95.2%, 93.7%, 97.5% and 96% for conventional ELISA, while these paramters for fast-ELISA were respectively as follows: 100%, 97.5%, 96.7%, 100% and 98.8%. Regarding standard-ELISA 3microg/ml of antigen, serum dilution of 1:500, conjugate dilution of 1:3000 and 30 min incubation were found optimal, while for fast-ELISA 3microg/ml of antigen, serum dilution of 1:125, conjugate dilution of 1:1000 and 5 min incubation were utilized. The present study indicates that fast ELISA can easily be performed in place of the standard ELISA for the serodiagnosis of human hydatidosis with the advantage of minimising consumed time and manpower hours. Moreover, this test can be utilized in screening tests to diagnos human hydatidosis

Evaluation of the enzyme-linked immuno-electro transfer blot [EITB] technique using hydatid cyst antigens B/5 and total IgG antibodies in laboratory diagnosis of human hydatidosis

To evaluate the validity of the Enzyme-linked Immuno-electro Transfer Blot [EITB] technique to diagnose human hydatidosis using sheep hydatid fluid antigens and human sera infected with hydatidosis. After preparing parasite antigen from sheep hydatid cyst fluid, all collected human sera infected with hydatidosis and other parasitic diseases as well as normal individuals, were analyzed by EITB test to evaluate its validity in diagnosing of hydatidosis. Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Iran. Seventy patients infected with hydatidosis confirmed by surgery, 15 with fasciolosis, 10 with toxocariasis, 15 with strongyloidiasis, 5 with amoebiasis, 10 with trichostrongylosis and 30 normal controls. The sensitivity, specificity, positive and negative predictive values of EITB test. Using total IgG antibody isotype, the sensitivity, specificity, positive and negative predictive values were 95%, 88.5%, 86.4%, 95.5% 97.3%, 64.8%, 60.3% and 97.7% for antigens Band5 respectively. The total IgG antibodies in hydatidosis patients documented the parasite AgB subunits i.e 12, 16, 23 kDa, also larger subunit of Ag5, namely 39 kDa. The study showed that although, EITB method was a time consuming test, but due to high validity could be considered as an authentic technique, especially when the diagnosis is vague and time is not imperative. Vic- Chancellery for Research, Tehran University of Medical Sciences, Iran. No Conflicts of interest exists

Evaluation of enzyme-linked immunosorbaent assay, using somatic and excretory-secretory antigens of strongyloides stercoralis for the serodiagnosis of strongyloidosis

Strongyloidiasis, caused by a nematode parasite so-called Strongyloides stercoralis is one of the major human intestinal nematode infections. Considering that stool examination for Strongyloides larvae is not a sensitive method and immunodiagnostic methods are more applicable for this purpose, so the present study was conducted to compare the somatic [S] and excretory - secretory [ES] antigens of Strongyloides stercoralis in IgG-ELISA to diagnose human strongyloidiasis. Serum samples obtained from 50 individuals infected with Strongyloides stercoralis. Sera from healthy control individuals, not infected with any parasitic diseases [n=/30] and from others with different parasitic infections including hydatidosis [n=20], toxocariosis [n=18], ascariasis [n=2], trichostrongylosis [n=10], and hymenolepiasis [n=2] were examined as well. The cut-off point for [S] and ES was 0.48 and 0.36, respectively. Thirty eight and 42 out of 50 individuals infected with Strongyloides stercoralis were also seropositive using [S] and ES antigens, in that order, whereas 15 cases of false positive reactions for [S] and 10 for ES antigen were detected when non-strongyloidiasis sera were examined, therefore the sensitivity of the test was 80.6% and 86.2% for [S] and ES antigens, respectively. The specificity of those antigens was calculated as 84.2% and 88.2%, correspondingly. It was concluded that overall ES antigen showed a more convincing diagnosis in comparison with [S] antigen, although every interpretation of the results should be in accompany with clinical manifestations and a history of the disease

Study on glutathione S-tranferase [GST] inhibition assay by triclabendazole.1: Protoscoleces [Hydatid cyst; Echinococcus granulosus] and sheep liver tissue

Hydatid disease is a term used to refer infection with the methacestode of Echinococcus granulosus parasite in humans, and echinococcusis is restricted to infection with the adult stage in carnivores.Glutathione S-Transferase [GST] represents the major class of detoxification enzymes from helminth parasites such as Echinococcus protoscoleces [PSC] and it is candidate for chemotherapeutic and vaccine design. Therefore, GST of protoscoleces could be a target for evaluation of drug effect as triclabendazole in hydatid cyst. For this purpose, GST enzymes were purified from protoscoleces of hydatid cyst and sheep liver tissue by glutathione affinity chromatography using a wash-batch method and subsequently detected their SDS-PAGE pattern. Afterward, GST specific activity levels were assayed in the whole extract and purified solutions spectrophotometrically at 30 C with reduced glutathione [GSH] and 1-chloro-2, 4-dinitrobenzen [CDNB] substrate. Finally, GST inhibition assay was investigated in the solutions by powder and bolus of triclabendazole. GST fraction as a 26 kDa [MW] band was obtained on SDS-PAGE. The level of GST specific activity in purified solutions was detected 10.24 mmol/min/mg proteins for protoscoleces and 37.84 mmol/min/mg protein for liver tissue. Comparison of the effect of powder and bolus of triclabendazole in solutions revealed inhibition concentration [IC50] 8.71 and 11.16 mg/ml for protoscoleces GST and 8.65 and 9.70 mg/ml for liver tissue GSTs, respectively. These findings suggest the possibility of selective inhibition of protoscoleces. GSTs by triclabendazol in vitro and use of these results for understanding of its molecular effect in vivo