Salmonella is one of the authentic bacteria which cause illnesses, may exist in raw material and food. The existence of these bacteria in food not only causes illnesses, but it also causes the downfall of production quality and reduction of economic growth of the area and country. In this study, 150 bulk raw milk samples were examined to comparison of PCR and conventional culture for the identification of Salmonella in raw milk. Firstly raw milk was cultured and examined through the conventional method; afterwards its supplementary procedures for isolating Salmonella were carried out. Regarding to the results of the culture method, six suspicious isolates were selected to carry out by PCR using invA gene. The results showed that none of the isolates were salmonella. Secondly DNA extracted from raw milk and samples were assessed utilizing the invA gene by PCR method. Regarding to the results 3 out of 150 examined samples were positive. Totally 2 percent of all samples were contaminated with Salmonella. The results of this study revealed that PCR is more potent than conventional culture methods to identification of salmonella in raw milk.

The NorA efflux pump considered as one of the contributors to antibiotic resistance in Staphylococcus aureus strains. One of the challenges of the researchers is finding natural plant compounds with the ability to inhibit the pumps. The aim of this study was to investigate the inhibitory effect of thyme (Thymus vulgaris) extract on NorA efflux pump in ciprofloxacin-resistant strains of S. aureus. Here, by using polymerase chain reaction (PCR), the presence of norA efflux pump was identified in 10 clinical and standard ciprofloxacin-resistant strains of S. aureus. The extract of thyme (T. vulgaris) was prepared using ethanol solvent and the effect of the extract against norA efflux pump was investigated by ethidium bromide method. In the following, after exposure to sub minimum inhibitory concentration of the extract, norA gene expression was evaluated using real-time polymerase chain reaction. Finally, we used the gas chromatography mass spectrometry (GC-mass) method to characterize the compounds with antibacterial properties. The results of PCR showed that all strains had a norA efflux pump, and ethidium bromide phenotypic method indicated that thyme extract had an inhibitory effect on all resistant strains with norA efflux pump. The norA gene expression in ciprofloxacin-resistant strains also decreased in sub minimum inhibitory concentration of the extract. By using gas chromatography- mass spectrometric, five chemical compounds of thymol, carvacrol, indole, 2-methoxy-4-methylphenol and quinic acid were determined as the dominant compounds of thyme (T. vulgaris) extract. Due to the antiefflux pump effect of thyme extract, it seems that this plant extract can be used as a good antibacterial component in the pharmaceutical industry of Iran.

Yersinia pestis, the causative agent of the zoonotic plague infection, is a major public health concern both as a threat and potential bioweapon. The objective of the present study was to establish a uniplex and multiplex - polymerase chain reaction (PCR) test for the specific detection of Y. pestis. PCR reactions performed by three pair primers which targeted the caf and pla genes located on the pFra and pPst plasmids and the irp chromosomal gene located on the ‘pathogenicity island’. After TA cloning of the PCR products, the test’s limit of detection (LOD) was determined. For evaluating the specificity, PCR reactions were performed with negative control bacteria. Assays were performed with the genome of Y. pestis which produced three DNA fragments of the expected sizes 00, 400 and 50 bp which corresponded to the irp, caf and pla genes, respectively. The lower LoD was 70 copy numbers for the caf gene and for the pla gene. In PCR reactions that used negative control bacteria, detectable fragments were not observed. Our method clearly discriminated Y. pestis DNA. The rapidity, specificity and sensitivity of this procedure suggest that it can serve as a useful alternative method for the inoculation of laboratory animals or the use of specific culture media for routine plaque surveillance and outbreak investigations. Another vital result of this study was the establishment of Y. pestis molecular detection technique in Iran.

Two structural antigens, haemagglutinin (HA) and neuraminidase (NA), are attractive candidates for the development of a genetically engineered vaccine against influenza. Recombinant vaccines are produced by a simple and effective method, although expected to induce an immune response to a specific antigen, remain to be further improved for their high effectiveness. On the other hand, a potent and effective vaccine against influenza should be able to induce both humoral and cellular immune responses. In the present study, the NA gene, which is more stable than the HA one, was amplified by polymerase chain reaction (PCR), and the band appeared in the range of 1400 bp. Then cloned into a eukaryotic expression vector pFastBac HTA. The purity of the expressed NA protein was analyzed on SDS-PAGE electrophoresis. In the western blot, a band with a weight of about 41 kDa produced by the recombinant baculovirus containing NA is observed compared to the uninfected cells used as a negative control, and a recombinant neuraminidase is confirmed. Finally, neuraminidase (NA) gene clone can be used to make recombinant human influenza vaccine.

Food contamination with fungi and the production of mycotoxins, such as aflatoxin, allow the toxins to enter human body. Continuous contamination with low doses of these agents can act as a major risk factor for hepatocellular carcinoma. Thus the present study was carried out to evaluate the detection of contamination in eggs with aflatoxin by PCR method. In a cross-sectional study, a total of 144 suspicious and 211 intake eggs were collected and three samples of fungi including aspergillus niger, penicillium expansum, and fusarium verticillioides as negative controls and 14 samples of aspergillus flavus as positive controls were selected and examined using TLC and PCR. The results were analyzed through SPSS software. By PCR, neither aflR, omt-A, and ver-1, nor-1 was detected in intake eggs by PCR. Of the suspected eggs, four samples with nor-1, two samples with aflR, and two samples with omt-A could be detected. Three samples of the 14 strains of aspergillus flavus were shown to be positive through the use of TLC and the four primers. One strain of aspergillus flavus was positive with all of the four primers; however, it was negative in TLC. The findings of this study indicated that PCR is a sensitive, fast, and specialized technique, but it cannot detect the presence of the fungi before the appearance of colonization. Thus for indicating toxcification, other complementary tests are also required.

Brucellosis is an important zoonotic disease in humans and animals. Brucellosis has been reported from most countries، but in some countries including Iran the disease is endemic. Consumption of milk and products of infected animals is a major transmission source of infection to humans. Since in most provinces of Iran such as Kurdestan province، cattle and sheep are kept close together، it is possible that animals are infected with non-specific Brucella species. The aim of this study was to use PCR in detection of Brucella species in milk، and also to detect the host specificity of Brucella abortus and Brucella melitensis in cattle and sheep. In this research، 60 milk samples from suspected cattle and 50 milk samples from suspected sheep in different villages of Kurdestan province were collected. DNA was extracted from all milk samples directly. In order to detect the Brucella spp. PCR was carried out using B4 and B5 primers on all extracted DNAs and in order to detect the B. abortus and B. melitensis، PCR was carried out with B. a، B. m and IS711 primers on all DNAs. Using PCR for detection of Brucella spp. 20 and 22 of milk samples were positive in cattle and sheep samples respectively. Using PCR، out of the 20 cattle positive samples، 9 samples were identified as B. abortus (biovar 1، 2 and 4)، and 2 samples were identified as B. melitensis. Also out of the 22 sheep positive samples، 15 samples were identified as B. melitensis and 1 as B. abortus (biovar 1، 2 and 4). Regarding the fact that in two milk samples of cattle، B. melitensis and in 1 milk sample of sheep، B. abortus were detected، the animals that were kept in close contact with each other may lose the host -specificity. Based on the results of this research، it is recommended that in order to detect Brucella spp. in milk samples. The vaccination status should be detected and using specific primers for detection of all biovars of Brucella abortus and Brucella melitensis، PCR and cultural methods should be carry out.

Salmonella enterica serotype Typhi is the causative pathogen of typhoid fever. Morbidity and mortality rates of the disease are 16×106 and 6×105 cases per year, respectively. Proper diagnosis of the disease and its suitable treatment have critical roles in morbidity and mortality reduction, especially in the developing countries. The traditional methods for detection of the organism are time consuming with low sensitivity. The aim of the present study is to design a new polymerase chain reaction (PCR) method for rapid detection of the organism.

Target specific viaB genes were used for primer designing using Generunner software. PCR tests were sat up using the standard Salmonella typhi genome. Besides, specificity of the method was determined using negative control bacterial genomes. For construction of positive control, determination of sensitivity of the method and limit of the detection, PCR products were cloned in a pTZ57RT plasmid vector.

The agarose gel electrophoresis of PCR products showed 441 bands for the viaB genes. PCR tests for control negative genomes did not create any band on the agarose gel. Results of PCR, enzymatic digestion and sequencing confirmed the cloning process and the positive control construct.

Considering the disadvantages of the classic methods for detection of the organism and the advantages of the molecular methods, the diagnostic kit proposes a suitable tool for rapid detection of the Salmonella typhi.

Septoria tritici blotch (STB), caused by Mycosphaerella graminicola (anamorph Septoria tritici), is one of the most important diseases of wheat (Triticum aestivum) world wide. planting resistant cultivars is the best method to manage STB environmentally. Many plant genes are induced or repressed and many others are up regulated or down-regulated to response a pathogen. In this study, Real time PCR were used to analysis of expression patterns of five genes involved in septoria leaf blotch resistance in wheat. After inoculation of resistant (Wang shui bai) and susceptible (Falat) cultivars with M. graminicola fungus, expression of these genes were investigated at 11 time points in 3 replications and the expression patterns of them were determined as relative to 18s reference gene. Comparision of these sequences using NCBI data base showed that these sequences had homology to neomenthol dehydrogenase, Lucine rich repeat protein kinase, natural resistance-associated macrophage protein, alpha 1,4-glucan phosphorylase and Tubby-like F-box protein. The peak expression for 1,4-glucan phosphorylase and Lucine rich repeat protein kinase were at 3 hours after inoculation and for neomenthol dehydrogenase the peak is at 12h. For natural resistance-associated macrophage protein the peak was at 3h but it was decreased and then increased again at second day. Tubby-like F-box protein was induced at 3h and had significant expression in this time but the peak expression for this gene was at 3 day after inoculation and we can call this pattern, a bimodal one. Some of these genes were induced early after inoculation and their expression increased significantly, but some others showed an induced expression several days after inoculation.

Q fever is a zoonosis disease caused by an obligate intracellular organism Coxiella burnetii. Raw milk or dairy products that are produced by unpasteurized milk may contain virulent Coxiella burnetii. The objective of this study was to determine the prevalence rate of C. burnetii in bulk milk samples from dairy sheep herds in Fars، Ghom، Kerman، Khuzestan and Yazd provinces، Iran. In the present study، 183 bulk milk samples from 59 dairy sheep herds were tested for C. burnetii using a nested PCR assay. The animals which their milk samples collected for this study were clinically healthy. In total، 3 of 183 (1. 6%) ovine bulk milk samples were positive; the positive samples originated from 2 of 21 (9. 5%) sheep breeding farms in Yazd. All bulk milk samples collected in Fars، Khuzestan، Ghom and Kerman were negative. Although no extensive prevalence study was undertaken، the results of this study indicate that clinically healthy dairy sheep in are important sources of C. burnetii infection. In conclusion، the results of the present study showed the importance of periodically monitoring the prevalence rate of C. burnetii in milk samples from dairy sheep herds in different provinces، Iran.

Methicillin-resistant Staphylococcus aureus has been considered as one of the pathogens involved in nosocomial infections. The study of drug resistance and typing of this bacterium is an effective measure to prevent and spread it. This study aimed to detect the mecA gene as well as SCCmec typing of Staphylococcus aureus isolates isolated from clinical specimens.

After collection, 308 clinical specimens were identified as Staphylococcus aureus isolates by confirmatory tests. Drug susceptibility was assessed by agar screen method and after DNA extraction by PCR, strains with mecA gene were identified and SCCmec typing was performed by multiplex PCR.

Staphylococcus aureus isolates showed resistance to methicillin in the agar screen method, but in the PCR method, 73% of them showed mecA positive gene. The results showed that these six strains were resistant to methicillin in the agar screen test but did not have the mecA gene in the PCR method. The highest percentage of samples belonged to SCCmec IV type (37%) and then belonged to SCCmecIII (31%).

More than 73% of Staphylococcus aureus strains had the mecA gene and 56% of them were registered as hospital-acquired strains.