Sequence analysis of the second internal transcribed spacer [ITS2] region of rDNA for species identification of trichostrongylus nematodes isolated from domestic livestock in Iran

Infectivity of herbivores with Trichostrongylus nematodes is widespread in many countries, having a major economic impact on breeding, survivability, and productivity of domestic livestock. This study was carried out on Trichostrongylus species isolated from domestic livestock in order to develop an easy-to-perform method for species identification. Trichostrongylus isolates were collected from sheep, goat, cattle, and buffaloes in Khuzestan Province, southwest Iran. Primary species identification was carried out based on morphological characterization of male worms. PCR amplification of ITS2-rDNA region was performed on genomic DNA and the products were sequenced. Phylogenetic analysis of the nucleotide sequence data was conducted employing Bayesian Inference approach. Consequently, a restriction fragment length polymorphism [RFLP] profile was designed to differentiate Trichostrongylus species. A consensus sequence of 238 nucleotides was deposited in the GenBank for Iranian isolates of Trichostrongylus species including T. colubriformis, T. capricola, T. probolurus and T. vitrinus. The designated RFLP using restriction enzyme TasI could readily differentiate among species having different ITS2 sequence. The molecular analysis was in concordance with morphological findings. Phylogenetic analysis indicated a close relationship among the sequences obtained in this study and reference sequence of relevant species. ITS2-RFLP with TasI is recommended for molecular differentiation of common Trichostrongylus species

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

Fatal case of Plasmodium vivax malaria in a splenectomized patient

Malaria is a major problem in tropical and sub-tropical countries, with high morbidity and mortality. Splenectomy makes patients more susceptible to serious bacterial and parasitic infections. We report for the first time in Iran a fatal case of Plasmodium vivax malaria, confirmed by microscopic and molecular [Semi-nested multiplex PCR] tests in a patient who had undergone splenectomy due to hemolytic anemia

Redefining Ph.D. education in Iran: a systematic review of literature

Reviewing the scientific literature concerning the analysis of different aspects of PhD education systems can help improve the knowledge promotion system in Iran. Different PhD training models around the world and their advantages and disadvantages have been investigated. The study explored the published papers using a systematic approach. Promotion plans in other countries were also investigated. Main databanks between 1998 and 2008 were systematically reviewed using ten standard and sensitive keywords. In the next step, the contents of eligible papers were analyzed and classified. Thirty-two eligible papers were included in the final analysis. The main themes in these papers were about the student admission, supervision of PhD students and evaluation methods. In addition, these papers discussed how universities might improve the links between their PhD students and the community and industry. Moreover, we found information about the variety of training method between different countries, the trends in the number of PhD students and their age, sex and ethnic compositions. Major challenges for Ph.D education include: discipline, law and order in Ph.D programs, quality of supervision, completion time, drop-out rates, increasing number of students, preparation for employment, labor market qualifications and skills, cost of education, inter- university student exchange programs and foreign students admission. Based on our findings, we need to pay more attention to diversify PhD training schemes particularly research-based PhD and professional PhD and to provide PhD education systems appropriate to the present and future needs of our society

[ correlation between clinical signs and genotypes of giardia duodenalis isolated from patients with giardiasis in Kerman City]

Giardiasis is one of the human parasitic diseases caused by a flagellate protozoan named Giardia duodenalis [G.lamblia]. Giardia is one of the most common organisms causing diarrhea in human and also a common gastrointestinal parasite in vertebrates. A total of 352 stool samples were collected from patients infected with giardiasis referred to health centers in Kerman city. Samples were examined by formalin- ether concentration procedure. First, DNA extraction was performed on 30 stool samples containing adequate Giardia cysts and then PCR-RFLP was done on glultamate dehydrogenase [gdh] marker. Clinical signs of patients were recorded in a questionnaire and their relationships with molecular results were analyzed. The highest rate of infection was in the age group of 0-12 years with significant difference with other age groups [P<0.0001]. The most common clinical signs were abdominal pain [71.7%], diarrhea [69%], abdominal cramping [54.1%] and the least common signs were malaise [20.4%] and fever [16.1%]. Of all 30 isolates, 18 samples [60%] were found as genotype All, 5 ones [16.7%] belonged to Al assemblage and 7 samples [23.3%] were BIII assemblage. There was a significant difference between genotyping of Giardia and clinical signs of diarrhea, abdominal signs and nausea [P<0.05]. Higher prevalence of Giardiasis was found in the age group below 12 years, but clinical signs in different age groups and two sexes were identical. Assemblage A showed correlation with mild intermittent diarrhea and assemblage B had correlation with persistent diarrhea

Spatial distribution and molecular identification of leishmania species from endemic foci of south-eastern Iran

Cutaneous leishmaniasis constitutes a major public health problem in many parts of the world including Iran. The primary objective of this study was to identify Leishmania species in endemic districts of Kerman Province, south-eastern Iran. This study was conducted by random sampling as cross- sectional descriptive between 2008 and 2010. Overall, 203 skin scraping smears were taken from the patients. Nested -PCR was performed to amplify variable minicircle fragments of Leishmania kDNA. Bam was the most infected district [71.1%], followed by Kerman [14.7%], Jiroft [5.4%], Baft [2.7%], Sirjan [1.6%], Shahr-e Babak [1.5%] and others [3.0%]. L. tropica was the most common species identified [194 cases, 95.6%], while L. major was found in only 9 cases [4.4%]. Of 203 identified patients, all species in Bam [l07 cases], Kerman [32 cases], Jiroft [l6 cases] and Shahr-e- Babak [l1 cases] were detected as L. tropica, whereas infected subjects in Baft and Sirjan showed L. tropica or L. major. Characterization of Leishmania species resulted in generation of 750 bp and 560 bp fragments, corresponding to those of L. tropica and L. major, respectively. L. tropica is the main species [95.6%] caused ACL in endemic areas of Kerman Province; however L. major is present in low level [4.4%]

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

Comparison of five simple methods for DNA extraction from Echinococcus granulosus protoscoleces for PCR-amplification of ribosomal DNA

Cystic hydatid disease is an important zoonosis, affecting humans and animals and is a significant public health and economic problem throughout the world and Iran. Since extraction of DNA from the parasite is a primary and crucial step which has a principal effect on PCR results, in the current study five simple methods for DNA extraction from protoscoleces of Echinococcus granulosus were applied and compared with each other. After collecting hydatid cysts from an abattoir, DNA samples were extracted from two cyst isolates from sheep, two from goats and two from camels using five different methods involving the use of glass beads, mechanical grinder, freeze-thaw, boiling and crushing. For all DNA samples extracted, one PCR assay based on amplifying rDNA-ITS1 region was performed and amplicons resolved on 1.5% agarose gels. The methods were compared regarding to DNA and PCR bands, time and cost effectiveness and laborious amount. The target DNA was successfully amplified from all samples using all methods produced an expected band size. All methods showed some advantages and disadvantages in PCR gels. The boiling method, which was the most time and cost effectiveness method, achieved the thickest bands in the PCR following grinder, crushing, freeze-thaw and glass beads. Boiling and crushing methods were the most suitable methods regarding their amplicon quality, easiness, quickness and cost effectiveness

Epidemiology of cryptosporidium infection of cattle in kerman/Iran and molecular genotyping of some isolates

Cryptosporidiosis is one of the most important parasitic zoonoses of human and animals. This infection is common in mammals and caused by the coccidian parasites of the genus Cryptosporidium. The Present study was designed to determine the epidemiology of Cryptosporidium infection in cattle in Kerman by using conventional morphological as well as molecular methods for molecular characterization. Fecal samples of cattle were collected fresh and directly from the rectum. Cryptosporidium oocysts were isolated by using formalin-ether sedimentation method followed by modified Ziehl-Neelsen staining technique. DNA of a number of isolates was extracted using QIAamp DNA stool mini kit [Qiagen]. A nested PCR-RFLP protocol amplifying - 850 bp fragment of SSU-rRNA gene used to differentiate species and genotypes of the isolates, using Sspl and Vspl as two restriction endonucleases. For each slide at least 20 oocysts were measured. Seventy eight of 412 cattle [18.9%] were found to be infected. Cryptosporidium infection was associated with diarrhea [P = 0.026] in a way that 31.8% of diarrheic cattle [14.44] and 17.4% of non diarrheic cattle [64.368] were infected. The rate of infection in suckling calves <2 months age was significantly higher than others [45.134 vs. 33.6%, P = 0.000]. In this study 4 isolates of C. andersoni and 8 isolates of C. parvum were found for the first time in Iran by using molecular techniques. Cryptosporidium infection is common in cattle of Kerman. Moreover, in spite of the presence of C.parvum as the dominant species in Iran, the presence of C. andersoni in Iran is reported for the first time by molecular techniques. Economic and public health problems resulted from infection by C.andersoni require more investigations in other parts of Kerman province and Iran

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

In vitro evaluation the effect of 1, 3, 4, thiadiazole new derivatives against

The main therapeutic compounds available against Leishmaniasis disease is pentavalent antimonyfcg compounds i.e. Glucantime and Pentostam. New antileishmanial compound is needed due to the emerge of drug-resistant leishmania agents in recent years. In the present study the antileishmanial activity of new 1, 3, 4 thiadiazole derivatives were evaluated. Promastigote stages of the parasites were cultured in RPM1-1640 containing 10% FBS, 100 lU/ml penicillin and 100 micro g/ml streptomycin. Mouse peritoneal exudate macrophages [MPEM] isolated from the peritoneal cavity of BALB/c mice were used and the macrophages were counted and the cell suspension was adjusted to 5X10[5] cell/ml. Macrophage monolayers in 8-well chamber slides were infected with stationary phase promastigote, at a 5:1 parasite/cell proportion and incubated at 37°C and 5% CO[2]. Serial dilution of thiadiazole compounds and tartar emetic as the control was added to the slide chambers and parasite survival index [PSI] was measured after 5 days. The Thiazolyl blue reduction [MTT test] was used to determine the antileishmanial effect of the compounds on extra celluto forms of the parasite and after 72 h. The CD's were read by 96-well scanner and IC[50] were calculated. Two thiadiazole compounds showed 6-67% antileishmanial activity in 4.6 micro g/ml concentration against intracellular forms of the parasites and also in MTT assay IC[50] of 3.6 -7.6 micro g/ml was determined. Due to high antileishmanial activity of some compounds, further studies on structure and activity of these compounds and new highly active derivatives is expected