Application of Multiplex PCR for Detection and Differentiation of Entamoeba histolytica,Entamoeba dispar and Entamoeba moshkovskii

Background

Entamoeba moshkovskii and E. dispar are impossible to differentiate microscopically from the pathogenic species E. histolytica. Multiplex polymerase chain reaction (Multiplex PCR) is a widespread molecular biology technique for amplification of multiple targets in a single PCR experiment.

Methods

For detection and differentiation of the three-microscopy indistinguishable Entamoeba species in human, multiplex PCR assay using different DNA extraction methods was studied. A conserved forward primer was derived from the middle of the small-subunit rRNA gene, and reverse primers were designed from signature sequences specific to each of these three Entamoeba species.

Results

A 166-bp PCR product with E. histolytica DNA, a 580-bp product with E. moshkovskii DNA and a 752-bp product with E. dispar DNA were generated in a single-round and multiplex PCR reaction.

Conclusion

We recommend this PCR assay as an accurate, rapid, and effective diagnostic method for the detection and discrimination of these three Entamoeba species in both routine diagnosis of amoebiasis and epidemiological surveys.     

Application of Mathematical Models in Combination with Monte Carlo Simulation for Prediction of Isoflurane Concentration in an Operation Room Theater

Applicability of two mathematical models in inhalation exposure prediction (well mixed room and near field-far field model) were validated against standard sampling method in one operation room for isoflurane. Ninety six air samples were collected from near and far field of the room and quantified by gas chromatography-flame ionization detector. Isoflurane concentration was also predicted by the models. Monte Carlo simulation was used to incorporate the role of parameters variability. The models relatively gave more conservative results than the measurements. There was no significant difference between the models and direct measurements results. There was no difference between the concentration prediction of well mixed room model and near field far field model. It suggests that the dispersion regime in room was close to well mixed situation. Direct sampling showed that the exposure in the same room for same type of operation could be up to 17 times variable which can be incorporated by Monte Carlo simulation. Mathematical models are valuable option for prediction of exposure in operation rooms. Our results also suggest that incorporating the role of parameters variability by conducting Monte Carlo simulation can enhance the strength of prediction in occupational hygiene decision making.     

The effect of a Pender’s health promotion model-based educational intervention on physical activity in office staff

Sport Sciences for Health - Physical inactivity is a common health issue and also a behavioral risk factor for a wide spectrum of defects. This study was conducted to investigate the effect of a...     

Angiotensin-converting enzyme gene polymorphism and the progression rate of focal segmental glomerulosclerosis in Iranian children

Aim: Focal segmental glomerulosclerosis (FSGS) is one of the most common forms of glomerulonephritis leading to end-stage renal disease (ESRD). A few clinical and paraclinical factors are considered as contributing factors in progression rate. However, there are controversial reports on the relationship between ACE gene polymorphism and rapidity of progression of FSGS to ESRD in different populations. To elucidate this issue, we investigated the relationship between the insertion (I) and deletion (D) ACE gene polymorphism and rapidity of progression of FSGS to ESRD in Iranian children. Methods: Forty-one children aged 1–18 years admitted to St AlZahra Hospital, Isfahan, and St Ali Asghar Hospital, Tehran, Iran, with idiopathic FSGS were enrolled. Renal death was defined as a glomerular filtration rate (GFR) of less than 50 mL/min per 1.73 m² or a decreased GFR to less than 50% compare to baseline. Reaching renal death in less or more than 2 years were labelled as rapid progressors (RP) or slow progressors (SP), respectively. Intron 16 of the ACE gene was amplified by the polymerase chain reaction technique. Results: Twenty-eight patients were male and 13 were female. In 15 RP patients, the genotype distribution was 26.6% DD, 6.7% II and 66.7% ID. In 26 SP patients, the genotype was similar (38.6% DD, 7.6% II and 53.8% ID, P > 0.05). There were no statistically significant differences for ACE I/D gene polymorphism between the two groups of patients (P > 0.05). Conclusion: Our study revealed no correlation between ACE I/D gene polymorphism and rapidity of progression of FSGS to ESRD in Iranian children.     

Clinical manifestations of chondromalacia patella in 260 Iranian patients

Background and aim: Chondromalacia patella, which is characterized by softening of the patellar cartilage, is the most common cause of anterior knee pain in young women. The aim of this study was to identify the clinical features of patients with chondromalacia patella in Iran. Methods: All patients under 40 years, complaining of mechanical knee pain who were referred to Amir A’lam Rheumatology Unit, with positive shrug sign and normal knee X‐rays during the period September 2000 to September 2002, were included in this study. After physical examination and knee radiography, patients with knee arthritis, knee osteoarthritis and knee periarthritis were excluded. Patients with the clinical diagnosis of chondromalacia patella were studied. The demographic data, clinical disease characteristics and disease course were recorded. Results: There were 260 patients. They were predominantly female (F : M, 2.6 : 1), in the third decade of life and a mean age of 22.8 years at the onset of disease. Bilateral involvement was found in 92% of patients. The first manifestation was knee pain. The history of trauma or swelling of the knee occurred in about 20% of cases. The history of dislocation was 3%. Sitting with flexed knees, stairs, and the use of Turkish WCs aggravated the knee pain. About one‐third had knee malalignment, mostly mild genu varus. Patella alta was seen in 1.6%. Q‐angle more than 15° was seen in 90.8%. Mean Q‐angle was 21.9°, mean patellar angle was 122.6°, and mean intercondylar angle was 141.5°. All patients had the shrug sign. About 90% had Rabot test and crepitation, 3.5% had knee effusion, and 1% had knee laxity. Lower limb discrepancy was seen in 6.2% and spinal deformity in 10%. Ninety‐three percent of the patients were treated by conservative (medical) therapy. Conclusion: So the classic case of chondromalacia patella is a woman in her third decade of life with mechanical knee pain and positive shrug test.     

First Molecular Identification of Sarcocystis ovicanis (Protozoa,Apicomplexa) in the Brain of Sheep in Iran

Background

The objective of the present study was to survey the presence of Sarcocystis in sheep’s brain in North Khorasan Province.

Methods

In general, 80 samples of sheep’s brain were collected from slaughtered sheep in slaughterhouses of North Khorasan Province. Tissue digestion method was used for observing bradyzoites in tissues. Histopathological processing tracing Sarcocystis and ensuing structural change in the brain tissue were conducted. PCR analysis was conducted on all the brain samples. Sequencing was done for one PCR product. Genotype was identified by Blast search and homology analysis.

Result

Sarcocystis spp. was found in one of the brain samples (1.25%) using tissue digestion method. The presence of bradyzoite was also confirmed in the prepared histopathological sections. PCR analysis was positive in one of samples. Genotyping of one sample proved that Sarcocystis species was Sarcocystis ovicanis and the nucleotide sequence of this parasite was deposited in the GenBank database under accession number No.{"type":"entrez-nucleotide","attrs":{"text":"KF489431","term_id":"540198437","term_text":"KF489431"}}KF489431.

Conclusion

Sarcocystis ovicanis can involve brain tissue of sheep and consequently causes clinical symptoms.     

P-glycoprotein A Gene Expression in Glucantime-Resistant and Sensitive Leishmania major (MRHO/IR/75/ER)

Background

Leishmaniasis is a parasitic disease caused by different species of Leishmania parasites with a wide range of clinical manifestations. Antimonial compounds such as meglumine antimoniate (glucantime) are the first line drugs for the treatment of leishmaniasis. However, according to reports of the drug resistance of parasites, the efficacy of antimonial compounds is low. The ATP-binding cassette (ABC) proteins are present in all organisms and mediate the transport of vital elements through biological membranes. One of the important mechanisms of resistance in Leishmania parasites is the overexpression of ABC efflux pumps. P-glycoprotein A (pgpA) is a related gene for ABC transporter in Leishmania species. The aim of this study was to compare the pgpA expression in laboratory-induced resistant L. major (MRHO/IR/75/ER) and sensitive parasites.

Methods

RNA extraction of promastigotes of sensitive and resistant clones was performed and total RNA was reverse transcribed. The real-time quantitative polymerase chain reaction (PCR) was used to assess RNA expression profiles and the expression levels were calculated using 2^-ΔCt method.

Results

The mean expression level of pgpA mRNA was 2.70 ± 0.51 in in sensitive Leishmania clone and 6.08 ± 1.50 in resistant Leishmania clone (P = 0.021).

Conclusion

The expression of pgpA gene in resistant strains of L. major was almost fivefold higher than those in susceptible strains. Therefore, this can be used in field isolates, i.e. overexpression of the gene can prove resistance in wild type field isolates.