محمدحسن شاه حسینی

As the world population grows, it is predicted that freshwater supplies will be rare in the 21st century. Despite abundant advances in water and wastewater treatment, waterborne diseases still threaten the health of the people of the world. Listeria monocytogenes bacterium is a pathogen that causes listeriosis. This pathogen can also cause meningitis, poisonous sepsis, and abortion in humans. One way of transmitting this microorganism is water and foodstuffs. Quick and accurate identification plays an important role in preventing infections. Also, due to the importance of Campylobacter jejuni in water and food industries and causing infection, toxication, and digestive problems in humans, the identification of this bacterium can be an effective step in preventing water contamination with Campylobacter jejuni. The aim of the present study was to identify Listeria monocytogenes and Campylobacter jejuni through culture and PCR and compare them in the water supply of Kermanshah city.

18 samples were collected from different water supplies of Kermanshah. DNA was extracted from standard Campylobacter jejuni and Listeria monocytogenes using a DNG-Plus kit. PCR reaction was optimized using specific primers. After determining the specificity and PCR detection limit, the collected water samples were examined and at the same time, the samples were cultured and examined.

From 18 samples of water supply sources in Kermanshah by PCR, Campylobacter jejuni was isolated from all samples, and Listeria monocytogenes was isolated from 17 samples, and also 4 cases of Campylobacter jejuni and 2 cases of Listeria monocytogenes were isolated by culture method. The results showed that PCR has a better performance than culture for detecting Listeria monocytogenes and Campylobacter jejuni.

 Cytomegalovirus (CMV) is a member of the herpesviridae family, which is a major cause of intrauterine viral infection. The aim of this study was to evaluate the diagnosis of CMV in women with idiopathic abortions by the polymerase chain reaction (PCR) technique.

 50 blood samples of women with idiopathic abortions were collected and viral genome was extracted using commercial kit DNG-Plus. The PCR technique was optimized using the standard CMV strain genome and then the Limit of Detection (LOD) and specificity of the test was evaluated using genomes other than CMV. The optimized PCR technique was performed on the samples and positive and negative controls and the results were analyzed by gel electrophoresis.

 using optimized PCR technique, 257bp product were observed in agarose gel. In the specificity test, positive results were obtained only with the CMV genome and a detection limit of 100 copies per reaction was obtained. Finally, out of 50 samples, two samples were positive for CMV.

 Idiopathic abortions are abortions that do not seem to have a specific cause. But obviously many factors including infectious agents that can cause abortion are difficult to identify them. One of these factors could be CMV and the results of this study showed that it is best to search and identify CMV in this type of abortion by molecular tests such as PCR, quickly.

King soldier bream is one of the species of family Sparidae, which is commercially and ecologically very important in the Iranian waters of the Persian Gulf and Oman Sea. The species of this family are morphologically very similar to each other and they may be mistakenly known as another species. Therefore, molecular identification of them has to be considered.

In this study, the phylogenetic relationship of king soldier bream fish based on mitochondrial region(CO1) was investigated using DNA sequencing method. For this purpose, 90 samples were collected from the soft tissue of the caudal fin using Ammonium acetate, and its quantity and quality were determined by Spectrophotometry and Electrophoresis. Optimal DNA samples for Polymerase Chain Reaction (PCR) and sequencing were examined. After gene amplification and sequencing, Bio Edit version 7.0.1, BLAST, MEGA 7.0.2 and DnaSP 5.10.01 software were used to draw phylogenetic trees.

Based on the phylogenetic tree obtained from the sequences, king soldier bream was divided into two main branches in Bandar Abbas, Bandar Bushehr and Bandar Chabahar areas. All samples of the first branch showed a sister relationship with the second branch and all the samples of branches one and two showed sisterhood with samples KJ012292 and kJ012291 from the Gene Bank in Italy with 82, 67 and 86% accuracy.

The results can provide useful information on the conservation and management programs, for the preservation, revitalization, and resource of the population this valuable species.

Herpes simplex virus type 2 (HSV-2) is one of the most important human pathogenic viruses. The virus causes genital herpes and can cause miscarriage, pregnancy complications, and infertility in women. The aim of this study was to evaluate the role of HSV-2 in idiopathic abortion by PCR.

Fifty blood serum samples of women with idiopathic miscarriages were collected and transferred to the laboratory. The PCR test was performed using standard HSV-2 strain DNA and then the limit of detection (LOD) and specificity of the test was determined using a different strain of DNA. A standard test was performed using positive and negative controls on the samples and the results were analyzed by agarose gel electrophoresis.

The PCR technique est optimized and 231 bp amplicon were observed in agarose gel. In specificity test only with DNA HSV-2 achieved positive results as well as a limit of detection 10 Copy / Reaction. Of the 50 samples studied, 9 (4.5%) were positive.

Idiopathic miscarriages are miscarriages that do not seem to have a definite cause. But clearly, several factors, including infectious agents that are difficult to identify, can cause miscarriage. One of these factors could be HSV-2. The results of this study showed that HSV-2 is one of the most important factors in this type of miscarriage and that HSV-2 can be quickly identified by tests such as PCR.

Leukemia begins with the destruction of hematopoietic bone marrow cells, and appears with the turmoil in the blood (immature blood cell proliferation, homeostasis disorder, degrees of inflammation, mutation, immunodeficiency, antioxidant deficiency, and infectious agents). Cytomegalovirus (CMV) as the largest member of the herpes family can cause latent infections for lifetime. Due to its high numbers in Iran, we decided to measure the level of CMV in leukemia by polymerase chain reaction (PCR) method, to quantify its prevalence.

100 blood serum samples of people with leukemia and 100 healthy controls were collected from hospitals in Tehran City, Iran, and DNA was extracted by phenol-chloroform method. </em>PCR test of CMV was optimized and evaluated for specificity and limit of detection (LOD). Optimized test was performed on all patient and healthy samples along with positive and negative controls. The PCR product was identified by agarose gel electrophoresis.

PCR test was optimized, and 257 bp product was observed in agarose gel electrophoresis. In specificity test, no product other than the CMV sample was observed. LOD was 100 copy/reaction. All healthy controls were negative, and 33% of the samples were CMV positive.

This study shows that CMV can be considered as an infectious and influencing factor in leukemia.

Various genetic and environmental factors are associated with multiple sclerosis (MS) and its progression. Environmental factors include infectious agents especially fungi. The immunological basis of MS and the effects of fungal infections on the immune system led us to compare the relative frequency of fungal infections in the serum of MS patients with controls.

One-hundred serum samples of patients with MS and control individuals were collected. After the DNA extraction, the specificity and sensitivity of the PCR test for universal fungal primers were investigated. Then, an optimal PCR test was performed on control and patient samples.

In this study, 575 base pairs (bp) of genes were considered as the target product using universal fungal primers. The PCR test was performed to check for the specificity with fungal, bacterial, and viral DNAs, along with fungal DNA replication and other reproductive DNAs. The positive cases were 11 and 4 among 100 patients and 100 healthy samples, respectively. Also, the detection limit in this reaction was 40 copies of DNA. The results using SPSS software and Fisher's exact test showed a significant difference between the two groups (P value= 0.052).

MS patients have a higher relative frequency of fungal infections due to weaker immune systems and immunosuppressive drugs. The presence of an active infectious agent in such patients can increase the duration of the destruction of the nervous system. Therefore, diagnosing the active infectious agent and prescribing an appropriate medication regimen can be considered as a contributing factor in preventing the progression of the disease. Therefore, more studies are needed on the types of effective fungal species and their pathogenesis mechanism.

In the past, morphological traits were used to classify organisms, but because of the problems encountered in classification using this method, the use of molecular markers was due to its advantages. Mitochondrial markers are one of the best options for the study of phylogenetic relationships. In this study, 10 specimens of green sea turtle were collected from three areas of Chabahar, Govater and Konarak in Sistan and Baluchestan province. After DNA extraction, the samples were sequenced. Sequences were studied using MEGA, BioEdit and Arlequin software. The results showed that based on the phylogenetic analysis of the studied population, all of the samples were placed in three populations. The average distance of the population with the help of P-distance was 0.18. The results of the Clustal W show that the greatest difference between the sequences is at the beginning and the end of the sequence. The amount of haploid variety was zero, and the number of polymorphic sites was zero, which indicates that the difference was not such as to make the species derivative. No recombination has taken place in sequences that are consistent with the nature of the area under study. The results of this study, can be used to study the genetic diversity of marine turtles in different geographical conditions. It can also help to manage this population and prevent extinction. It can also be a good solution to provide a suitable Gene bank for the identification of this species through molecular techniques, especially in cases where only the eggs or meat of this animal is present.

Polychaetes are ecologically important in the food chain, and also vital in ecosystem monitoring and environmental stress assessment. In the current study, sequence variation in a segment of COI gene sequencing by mitochondrial DNA barcoding and comparing nucleotide divergence was employed for molecular analysis of Perinereis polychaetes (Nereididae). Analysis of 50 sequenced specimens, representing 39 new sequences, and 11 sequences with 100 percent similarity with GenBank records, which revealed 1.8 times more sequence divergence between than within species (1.96 versus 3.52%). Phylogenetic analysis data indicated that maximum divergence within barcode clusters increased by the number of specimens. The phylogeny also suggests that most of the identified Perinereis species in this research showed low genetic relationship towards similar species from other world regions. Assessment of genetic similarity rate showed that Perinereis polychaetes in Bushehr province were highly similar, while Perinereis polychaetes in Bandar Abbas were less similar to Bushehr Perinereis polychaetes, the observed divergence was mostly at subspecies level and it can be concluded that in terms of species structure, Perinereis polychaetes of Deylam, Bushehr, Dayyer and Bandar Abbas, showed low genetic divergence.

Different environmental factors, such as infection have cause Alzheimer's disease. Studies have shown that Herpes simplex 1 and 2 and cytomegalovirus viruses have relations with Alzheimer disease. Human cytomegalovirus viruses (CMV) induces infection in central nervous system. It is need a study investigating the relation between different viruses with Alzheimer. The current study, investigates the relation between different viruses Alzheimer. In this study, 100 patients with Alzheimer were studied and blood samples were collected (1.5 ml per patient). Following extraction of viral DNA, the samples were investigated by PCR method. The data for peoples with positive PCR were analyzed by Kruska-Walis test of SPSS software (version 23). The prevalence of CMV, HSV2 and HSV1 was observed in 27, 8 and 4 people that means 27%, 8% and 4%. These results suggest that CMV infection is associated to increased risk of AD and a quick rate of cognitive decline in elderly populations. In sum, all the studied viruses were identified, but cytomegalovirus virus had higher frequency.

Bacterial infections of the genital system are common causes of infertility. One of the most important causes of male infertility is seminal and genital tract infections. In the meantime, a wide range of bacteria, in varying degrees, contribute to infertility in men. Helicobacter pylori may be involve in infertility. Because antibodies against this bacteria have been significantly detected in the genital fluids of infertile patients. The aim of this study was to determine the presence of Helicobacter pylori in the semen of the infertile men with rapid molecular PCR method and determine its percentage.

100 samples of semen collected from Saram hospital and DNA Extracted from these specimens using Boiling/DNG_PLUS method.  Optimized PCR test was performed on samples. PCR test evaluated from the respect sensitivity and specificity.

In this study, amplicon with the size of 294 bp, observed with agarose electrophoresis.  In the specificity test, primers created only a band for H.pylori DNA. Of the 100 samples tested, only 10 samples (10%) were positive for DNA of H.pylori.

According to the results of this study, H.pylori may be one of the bacterial agents that can be considered for male infertility, of course requires further studies. While PCR is suitable, quick and sensitive molecular technique for detection of H.pylori in infertile men.

Aim and Bacterial arthritis is one of the arthritis diseases known that can rapidly cause joint damage. Among the bacteria causing septic arthritis, Mycobacterium tuberculosis (MTB) is one of those that rarely produce arthritis. The aim of this study is to examine the presence of MTB in synovial fluid of patients with arthritis using Polymerase Chain Reaction (PCR)
This study was carried out on 70 synovial fluid samples gathered from Shariati Hospital. DNA was extracted using Phenol-Chloroform standard extraction technique. PCR test optimized on the basis of IS6110 target gene. Samples were analyzed by PCR test after evaluation of specificity and sensitivity of PCR.
PCR test was optimized and the 317-bp amplicon detected by 1.5% agarose gel electrophoresis. Limit of detection (LOD) was estimated as 100 copy/Reaction. MTB DNA was detected in 4 (5.7%) synovial fluid samples of patients with arthritis.
CONLUSION: First step in treatment is rapid and accurate diagnosis. MTB have some characteristics including slow generation making the identification difficult and exhausting through culturing and biochemical tests that sometimes leads to ambiguous results. Results of this study confirm that MTB could play a role in bacterial arthritis and rapid diagnosis using PCR provides us with accurate treatment.

Probiotics are microorganisms that if administered in adequate amounts would
have beneficial effects on the host's health. Bacillus coagulans is a probiotic bacteria which
sometimes mistakenly called Lactobacillus sporogenes. B. coagulans is a gram positive and
facultative aerobic and anaerobic, catalase positive, oxidase negative and endospore forming
bacteria. The aim of this study is the rapid molecular detection of B. coagulans in various
probiotic products. PCR test was optimized by using B. coagulans standard strain.
Limit of detection (LOD) and specificity of this test were investigated by using related species
of DNA. Samples of probiotic products which contain B. coagulans were collected from the
Iranian market and various methods of DNA extraction were optimized for these products. Amplicon with the size of 450bp was amplified by PCR test and observed by
electrophoresis method. In specificity test none were positive using DNA for intended
organisms. B. coagulans species were identified in all the samples by PCR method. According to the results of this study B. coagulans was detected in all samples.
It was concluded that PCR is a reliable and accurate method to identify these products that
might contain B. coagulans probiotics bactria.

Background and purpose: Ureaplasma urealyticum is one of the most common causes of Non-Gonococcal Urethritis (NGU). Asymptomatic clinical infection caused by this bacterium can cause malnutrition of the sexual attachment glands and its presence in semen contributes to lower fertility. The aim of this study was to identify Ureaplasma urealyticum in semen of infertile men using PCR method as an accurate diagnostic method. The PCR test was optimized by standard strain to detect Ureaplasma urealyticum and then was studied in terms of specificity and limit of detection (LOD). Semen samples were collected from 100 infertile men and each sample was divided into two parts: the first part was tested by semen analysis and the second part was tested by PCR method. DNA was extracted using phenol-chloroform method and the PCR test was done for detection of Ureaplasma urealyticum. Among the semen samples, 16 cases (16%) were found to be positive for Ureaplasma urealyticum. According to the spermogram test, the leukocyte level was also more than normal level in these samples (0-1 Mil/ml). Screening of infertile couples for Ureaplasma urealyticum infection in those without clinical symptoms, thereby performing timely antibiotic therapy play key roles in treatment of infertility. Hence, PCR method is introduced as a valuable and reliable technique to identify Ureaplasma urealyticum.

Haemophilus influenza is a human-restricted Gram-negative bacterium that is part of the normal nasopharyngeal flora of most humans. H. influenzae can cause diseases such as pneumonia, meningitis, bacteraemia, sinusitis and acute otitis media. Nonculture methods like PCR have become available during the last two decades, providing early and accurate diagnosis of bacterial diseases. The purpose of this study was to design a PCR assay for the rapid detection of H. influenzae bacterium. In this study the target gene was P6 so the specific primers were designed for it. The PCR reaction was set up on the DNA genomic of H. influenzae. To create a positive control, the PCR product was cloned in the pTZ57R/T vector and was transformed into E. coli JM107. The sensitivity of this assay was tested by a serial dilution of the positive control. A total of 72 clinical samples collected from patients with sinusitis were analyzed by PCR. PCR results showed a band of the expected size 273 bp. Sensitivity results indicated that the limit of detection of the assay was 1 CFU/ml. No amplification was observed after PCR with any of the microorganisms tested. There was 19 (26%) positive numbers of H. influenza in the samples in this study. The PCR method showed to be rapid, sensitive, highly specific, and cheaper than commercial methods. The PCR methods could be easily adopted by public laboratories of developing countries for diagnostic purposes.

H. Pylori is an important cause of human peptic ulcers and stomach cancers. Once the bacterium is placed in the water, it changes into a viable, but non-cultivable coccoid form, which is considered as an important factor to spread out the infection. The aim of this study was to evaluate the survival rate of coccoid forms of H. pylori in water samples, using PCR and culture methods. This experimental study was performed on 10 strains of H. pylori isolated from clinical specimens. Isolates were added to water at the temperatures of 4°C, 22°C, and 37°C and incubated at intervals of 1 and 2 months. Each time, the samples were cultured on Brucella blood Agar medium. Following DNA extraction, the presence of glmM gene was confirmed using PCR method. Furthermore, the sensitivity and specificity of PCR method were evaluated. While
culturing in first month at 4°C showed no H. pylori coccoid growth on medium, some positive results (growth rate of 10% and 20%, respectively) were detected at 22°C and 37°C during the same month. No positive result was obtained during the second month. Performing PCR, with more sensitivity as compared to culturing method, identified H. pylori coccoid growth of 10%, 30%, and 40%, for the first month, and 0%, 20%, and 30% for the second month at 4°C, 22°C, and 37°C, respectively. The results of this study showed that the non-cultivable cocoid forms of H. pylori in water can be detected by non-culturing methods such as PCR which is sensitive, specific, and accurate.

In this study, the effect of different concentrations of potassium sorbate and sodium benzoate (300 , 400, 500 ppm) on microbial characteristics (total count, mold and yeast) of dried sour cherry with moisture of 25 % was evaluated during 6 months storage at temperatures of 8, 22, 37 °C. Microbial characteristic of samples were evaluated in the 1th, 3th, 6th months. The results showed that the dried sour cherry which treated with potassium sorbate and sodium benzoate during the six months of storage in vacuum packages had no microbial spoilage. The results showed that by using sodium benzoate and potassium sorbate at optimum condition, the amount of mold and yeast decreased from 11.67×10 cfu/gr (in control dried sour cherry) to 3.67×10cfu/gr and 3.33×10 cfu/gr respectively. Also, by using sodium benzoate and potassium sorbate at optimum condition, the amount of total count decreased from 2.13×103 cfu/gr in control sample to 0.50×103 cfu/gr and 1.23×103 cfu/gr respectively. The amount of mold and yeast and total count were decreased by increasing concentrations of potassium sorbate and sodium benzoate. By increasing temperature (from 8 to 37°C ) and time(1 to 6th month) due to the decomposition of potassium sorbate and sodium benzoate, antimicrobial effects of these compounds significantly decreased (p

Aspergillosis is one of infectious disease that it causes difficulties for human and poultry. The main aim of this study is the diagnosis of Aspergillus parasiticus and flavus in forage samples by Duplex PCR method and comparison with culture and direct examination. After extracting DNAs of standard strains of Aspergillus parasitcus and flavus along with primers of each fungus and PCR testing, genes of each fungus were amplified and PCR product was cloned using TA cloning by pTZ57R plasmid. After optimizing Duplex PCR (D-PCR) monoplex PCR tests, 50 forage samples by Duplex PCR method, culture and direct were tested.In the optimized PCR test, 343bp and 413bp products of Aspergillus parasiticus and flavus were respectively amplified. Fifty samples of forage by Duplex PCR method, culture and direct were tested. 13 samples were positive only by Duplex -PCR and 15 samples by both tests direct and culturing were positive and 26 samples were negative with three methods. Results of McNemar's test is equal about the three methods tests for diagnosing Aspergillus parasiticus and flavus (p=0.824). Molecular methods such as D-PCR are faster techniques than direct testing and culturing for diagnosis of Aspergillus parasiticus and flavus but there arent difference between three methods for diagnosing Aspergillus parasiticus and flavus but with D-PCR we could diagnosis Aspergillus parasiticus.

Fusarium head blight (FHB), is the most destructive disease of wheat, producing the mycotoxin deoxynivalenol, a protein synthesis inhibitor, which is harmful to humans and livestock. dsRNAmycoviruses-infected-isolates of Fusariumgraminearum, showed changes in morphological and pathogenicity phenotypes including reduced virulence towards wheat and decreased production of trichothecene mycotoxin (deoxynivalenol: DON). Previous studies indicated that over expression of yeast acetyl transferase gene (ScAYT1) encoding a 3-O trichothecene acetyl transferase that converts deoxynivalenol to a less toxic acetylated form, leads to suppression of the deoxynivalenol sensitivity in pdr5 yeast mutants. To identify whether ScAYT1 over-expression in transgenic tobacco plants can deal with mycotoxin (deoxynivalenol) in fungal extract and studying the effect of dsRNA contamination on detoxification and resistance level, we have treated T1 AYT1 transgenic tobacco seedlings with complete extraction of normal F. graminearum isolate carrying dsRNA metabolites. First, we introduced AYT1into the model tobacco plants through Agrobacterium-mediated transformation in an attempt to detoxify deoxynivalenol. In vitro tests with extraction of dsRNA carrying and cured isolates of F. graminearum and 10 ppm of deoxynivalenol indicated variable resistance levels in transgenic plants.Discussion and The results of this study indicate that the transgene expression AYT1 and Fusarium infection to dsRNA can induce tolerance to deoxynivalenol, followed by increased resistance to Fusarium head blight disease of wheat.

Mycoplasma is one of the smallest alive microorganisms, having the ability of transmitting from microbiological filters makes it to be a severe and important contaminant for cell culture, biological and biotechnological products. Using a rapid and sensitive method for diagnosing Mycoplasma’s infection is the first priority. This method should be able to discover typical Mycoplasma’s infection, but for meeting the standard we need to produce the internal control competitively in Mycoplasma spp PCR recognizing, which is the main aim of the project. Using of special primers for IS6110 target, PCR test was optimized. Besides, the composite primers for IC- M. spp were designed then the PCR was optimized. The IC- M. spp which amplified in constringent condition, was ligated in pTZ57R plasmid, transformed in E.coli JM107 and was cloned. Specification, sensitivity of test and the number needed in each test. Also PCR with internal control were defermined. The number of IC, which is needed in each PCR reaction was scanned in the matter of dilution and PCR response with IC. PCR amplicon for Mycoplasma spp and IC- M. spp with special primers were 272bp and 660bp respectively, so there was a significant different between their size. Despite the high speed and accuracy of PCR, false positive and negative results, which are caused because of PCR inhibitors, are the important problems of this technic that can reduce its efficiency. Using another DNA as an internal control can detect these inhibitors. Indeed, amplification of this DNA shows correct amplification and detection steps.

Mycoplasma pneumoniae bacteria, is one of the most important factor in causing of respiratory infections. Serological and molecular detection methods have their own limitation. Due to this limitation, the application of these methods in all diagnostic laboratories is not possible. Therefore this study was done to determine the rapid detection of Mycoplasma pneumonia by loop mediated isothermal amplification (LAMP). In this descriptive laboratory study, nasopharynx samples were collected from 92 patients with atypical pneumonia. DNA sample were extracted by boiling method. Six specific primer pairs were designed for LAMP technique by primer explorer ver 4 software. LAMP product identified by adding SYBR Green. Limit of detection and specificity tests have been done for optimizing LAMP test and optimized test carry out for each sample. The LAMP test was optimized using the large Bst enzyme fragment at 66 degree temperature for 1 hour. The detection limit of the test obtained 1 CFU and the DNA replication does not observed in non of the examined pathogenic factors. Out of 92 clinical samples using LAMP technique, 73 cases were negative (80%) and 19 cases were positive (20%). The loop-mediated isothermal amplification technique is simple, convenient and available method for detection of Mycoplasma pneumoniae.

Leptospirosis caused by infection with pathogenic leptospires، which is the most prevalent zoonotic disease in the world. The outer membrane proteins (OMPs) of pathogenic leptospires such as LipL41 play a crucial role in pathogenesis of this disease. Therefore a major challenge to develop an effective vaccine against leptospirosis is application of basic research on the OMPs of leptospires to improve vaccine development. The aim of this study was cloning and analyzing of the lipL41 gene from vaccinal serovars of leptospires in Iran، in order to identify genetic conservation of this gene. Three vaccinal serovars of Leptospira were used in this study. The lipL41 gene of these serovars were amplified and cloned in the pTZ57R/T vector. The recombinant clones were confirmed by colony-PCR and sequencing. The sequenced genes were analyzed for their homology between them and other submitted sequences in Genbank database using the BLAST and MegAlign program. PCR amplification of the lipL41 gene resulted in the 1065 bp gene product in vaccinal serovars tested. In our study، nucleotide sequencing results showed high similarity (>94%) within the leptospiral vaccinal serovars. The genetic conservation of the lipL41gene among different serovars of Leptospira indicated the capacity of utilization of this gene for development of recombinant vaccine against leptospirosis.

Obtaining information regarding pathogenesis and prevalence of extended spectrum beta-lactamase (ESBL) producing genes seems to be necessary، since it can promote prevention modalities and treatment of the infections caused by bacterias such as Klebsiella. The aim of this study was early identification of the blaTEM gene in Klebsiella، using polymerase chain reaction (PCR) technique. In this cross-sectional study، conducted form April to September 2013، 70 Klebsiella isolates were extracted from clinical samples (i. e.، wound، urine، sputum and blood) using biochemical tests، including non-state fermentation and triple sugar iron، negative indole، motile and methyl red، as well as positive Voges–Proskauer and urease tests. Subsequently، the frequency of ESBL producing strains was determined by means of combined disk method. DNA was extracted by boiling and was investigated for the presence of TEM gene using the PCR approach. In the 70 Klebsiella isolates، 11 cases of ESBL phenotype were observed، of which 10 cases contained TEM beta-lactamase resistance gene. In addition، 9 out of 59 samples (26%) of negative ESBL in antibiogram، were determined positive in terms of blaTEM gene using PCR method. Given the increasing prevalence of ESBL producing strains and poor diagnosis rate of antibiotic resistance through antibiogram method، applying more accurate techniques such as PCR is highly recommended.

Chronic sinusitis(CS) is one of the most prevalent chronic illnesses that affecting persons of all age groups. It is an inflammatory process that involves the paranasal sinuses. There isnt definitive and consistent data concerning the distribution of bacterial species in patients with Chronic Sinusitis. Pseudomonas aeruginosa has emerged as a potential pathogen in the immunocompromised patients, so this microorganism is one of the most important cause of Chronic sinusitis. The purpose of this study is molecular detection of sinusitis caused by P.aeruginosa. 50 specimens was provided from the secretion of maxillary and frontal sinuses of patients from Rasoule Akram hospital during operation. Genomic bacterial DNA was extracted by DNP kit and detection of this bacteria was proceeded by employing sequence-specific target namely the outer membrane protein (oprL) gene locus and designing primers. PCR optimized and sensitivity and specificity tests was performed. Amplicon was cloned by T/A Cloning method and was used for sequencing and positive control. The product of optimized PCR with 504 bp length correctly amplified and observed on electrophoreses gel 1.5% and confirmed by sequencing. Evaluation of the selected primers with 8 various DNA demonstrated 100% specificity. Sensitivity of optimized test was Evaluated 10 CFU of bacteria. From the 50 samples, 22% of specimens were positive for P.aeruginosa. This study indicates that molecular detection of P.aeruginosa employing the oprL gene target is a rapid and useful technique for detection of P.aeruginosa.

S. aureus, the second most common cause of nosocomial infections, is regarded as an important factor in the severe infections of the community. Methicillin-resistant strains of this bacterium involve a major pathogen which can cause disease and mortality in Iran and the world. Its treatment seems to be difficult due to the prevalence of resistance to most commonly-used antibiotics. In fact, methicillin -resistant strains need to be identified precisely and rapidly. Therefore, this study intended to assess the resistance to methicillin via the disk diffusion method and PCR for mecA gene. This study was conducted on 100 strains of Staphylococcus aureus collected from clinical various samples of Moheb and Milad hospitals in Tehran. Sensitivity to antibiotics was determined by the disk diffusion method and gene resistance (mecA) was examined by PCR method. Moreover, specific primers were explored and the results were compared. The prevalence of methicillin-resistant strains by the disk diffusion method was 50% (n=50), whereas 74% (n=74) were determined to have mecA gene via PCR analysis. Out of these 100 samples, 61 samples belonged to Moheb hospital, among which 47.54% (n=29) were observed to be methicillin-resistant by disk diffusion method and 60.65% (n=37) via PCR method. Other 39 samples were obtained from Milad Hospital, of which 84/53% (n=21) demonstrated resistance to methicillin via disk diffusion and 87/94% (n=37) via PCR method. The study findings revealed that due to high prevalence of methicillin resistance, a quick and detailed identification method of MRSA is required. Since disk diffusion method proposes relatively high false-negative results and is observed to have a time-consuming nature, PCR can be taken in to consideration as the best method in order to identify methicillin-resistant strains.

Aim and Fusarium solani, may occur as a mixed infection with HSV and due to similarities in their clinical features, sometimes it leads to misdiagnosis. The aim of this research was the designation and optimization of PCR technique for the diagnosis of F. solani as the main etiological agent or as a mixed infection in corneal samples suspected of having HSV 100 samples of DNA from clinical cases (Include 65 negative and 35 positive samples for HSV), suspected to herpetic keratitis were collected. The PCR test for detecting F.solani in these samples, with specific primers ffuso1 and rfuso2 and mt cytb as the target gene with 330 bp product was optimized. The specificity and sensitivity of the test were then evaluated. Among 65 negative samples for HSV, two samples were positive for Fusarium solani and the PCR 330 bp product were amplified in these two cases. The sensitivity of the test was one copy of Fusarium solani genome and specificity was 100%. The results of PCR test show that, the cases suspected to herpetic keratitis are caused by Fusarium solani., Accurate diagnosis of ethological agent in the cases of clinical features similarity or mix infection, is possible by molecular diagnosis.