Background and Aim: Culture and specific staining (including Zeil-Nelson and fluorescent methods) are standard measures for the diagnosis of tuberculosis (TB). Because these methods are time-consuming and, sometimes, due to their low accuracy faster and more accurate methods are necessitated. Methods, which can substitute invasive procedures, when obtaining smear samples and culture is not possible, and in addition to being simple and fast, they have an acceptable diagnostic accuracy.The aim of the present study was to verify the diagnostic value of blood Polymerase Chain Reaction (PCR) method in pulmonary TB.Materials and Methods: This case-control study included 64 proven pulmonary TB cases (according to The National TB Protocol) and 28 subjects who were completely healthy. 4.5ml of blood was derived from each participant and then mixed with 0.5ml EDTA. Finally, DNA extraction and PCR testing using SI 6110 primers was performed for all blood samples.Results: Mean age of the cases and controls was 49.8±18.6 and 48.2±18.5, respectively. 49.2% of the cases and 25% of controls were male. Blood PCR in 23 patients with TB was positive, but none of the controls had a positive PCR (thus, sensitivity of 35.7% and specificity of 100%).Conclusion: With regard to specificity of 100% in PCR method (despite its low sensitivity), in conditions where there is no access to an appropriate specimen, a positive blood PCR can obviate invasive procedures and rapid and definitive diagnosis of the disorder and timely treatment of the patient, his life is saved.

Background: The genitourinary system is one of he most common sites of infection in non-pulmonary tuberculosis (TB). The clinical symptoms and radiologic findings of urinary TB are nonspecific. Current diagnostic tests are of low sensitivity and labor-intensive. Therefore, this study was aimed to evaluate diagnostic value of urine PCR in genitourinary tuberculosis (GNTB).Methods: This was a descriptive study on 33 patients with confirmed genitourinary TB. Demographic data, clinical symptoms, laboratory and radiologic findings were collected. For each patient, three consecutive early morning urine specimens were examined by PCR. The diagnostic value of PCR in mycobacterium tuberculosis (MTB) in comparison with standard microbiological methods was assessed.Findings: There were 33 patients with a mean age of 47.27 16.1 years. The most common presenting symptoms were irritative voiding symptoms (51.5%), flanks pain (27.2%), gross hematuria (9%) and suprapubic pain (9%). Laboratory findings in U/A were hematuria (75.8%) and pyuria (60.6%). IVU was abnormal in 61.5% of patients. Most common abnormalities were pyelocalyceal dilation (44%), ureteral stricture and hydroureter (37%) and multiple small calyceal deformities (25%). Of the 33 patients PCR for MTB was positive in 16 cases (48.5%). In patients with abnormal IVU, PCR was positive in 62.5%.Conclusion: A high index of clinical suspicion is necessary for diagnosis of GUTB. PCR is recommended for instant diagnosis and screening before further examination, it cannot be the only method in identification of GUTB.

Background: Dermatophytes are a group of fungi that attack keratinous tissues of the skin, hair, and nail in humans and animals, and cause infections called dermatophytosis (tinea). Since identification of pathogenic fungi at the species level is essential for the detection of the source, control and prevention, and identifying epidemiology of infection, it is necessary to use specific and sensitive diagnostic methods to identify the causes of dermatophytosis. Methods: The clinical samples (skin, nail, and hair) of patients with dermatophytosis in Mashhad City, Iran, were cultured in Mycosyl Agar culture media, and the DNA of obtained dermatophyte colonies were extracted by specific kit. The internal transcribed spacer (ITS) gene was amplified and sequenced by ITS1, ITS4 primers. Finally, the sequencing results were analyzed using SeqMan software, and were compared with the data of the global genebank. Findings: In this study, 80 dermatophyte isolates were sequenced, which included 9 dermatophyte species as 23 (28. 8%) Trichophyton (T. ) interdigital, 18 (22. 5%) T. tunsorans, 10 (12. 5%) Epidermophyton fluccosum, 10 (12. 5%) of T. mentagrophytes, 8 (10%) Microsporum canis, 4 (5%) T. rubrum, 4 (5%) T. benhamiae, 2 (2. 5%) Nannizzia (N. ) fulvum, 1 (1. 2%) N. persicolor. Conclusion: According to report the rare species of dermatophytes in this study, the use of molecular methods such as sequencing of the ITS gene can determine the diversity of dermatophytes in a region more precisely than morphological methods.

Background: Disseminated candidiasis, diabetes mellitus, pregnancy, and consumption of widespread antibiotics are predisposing factors of urinary tract infections due to Candida species. The prevalence of nonalbicans species is increasing. The aim of the present study is to identify Candida spp. isolated from candiduria patients in Isfahan, Iran using Polymerase Chain Reaction-restriction fragment length polymorphism (PCR-RFLP) method.Methods: The genomic DNA was extracted by fast technology for analysis-card (FTA-card). Polymerase Chain Reaction (PCR) was used to amplify ITS1-5.8s-ITS2 region then PCR products were digested with MspI restriction enzyme. PCR and RFLP products were revealed on 1.5 and 2% agarose gel electrophoresis, respectively.Findings: Out of 3200 urine samples 80 samples with more than 104 CFU/ml were involved in this study. Patients including 4 (5%) were males and 76 patients (95%) were females. Predisposing factors were diabetes (47.5%), urinary tract infection (38.8%) and kidney stone (5.0%). The colony count of 46.3% of the urine samples were with 50´103-99´103 CFU/ml. Candida glabrata was the most prevalent species among isolates (41.3%).Conclusion: Increasing of non-albicans Candida species and drug resistance of isolates are main challenges of Candida infection treatment particularly in patients with diabetes and urinary infections. Identification of Candida species is inevitable for better management of infection.

Background: Klebsiella pneumoniae (K. pneumonia) is an opportunistic pathogen and one of the most common causes of nosocomial infections, some of which are now resistant to antibiotics and make infection difficult to treat. The main purpose of this study was to use multiplex Polymerase Chain Reaction (PCR) to detect tetracycline-resistance genes in clinical isolates of K. pneumonia. Methods: After collecting clinical samples from hospitals in Mashhad City, Iran, in 2021, K. pneumoniae was isolated and identified by biochemical and molecular methods (Kp16S rRNA specific primer). Antibioticresistance pattern was determined for 30 isolates by disk diffusion method. After designing primer and its blast, detection of tet genes was optimized by PCR. Finally, multiplex PCR was set up using three pairs of primers and positive control. Findings: The highest and lowest antibiotic resistance of the isolates was for ampicillin (90%) and ceftriaxone (26. 66%), respectively, and 53. 33% of Klebsiella isolates were phenotypically resistant to tetracycline. Multiplex PCR was set up by annealing temperature of 55°, C. Simultaneously with detection of Kp16S rRNA gene, tetA and tetB genes were detected in 87. 5% and 68. 75% of tetracycline-resistant Klebsiella isolates, respectively. 56. 25% of the isolates carried both tetA and tetB genes, simultaneously, and 12. 5% of the isolates transferred one of the tetA or tetB genes. Conclusion: The results show that the multiplex PCR technique designed in this study is a fast, accurate, and sensitive method for identifying antibiotic-resistant K. pneumoniae that can be used for clinical samples or in epidemiological studies.

What is PCR purification.PCR Purification Protocol is designed to purify double stranded DNA fragments from PCR and other enzymatic Reactions. Fragments from 100bp to 10kb are purified from primers, nucleotides, Polymerases and salts using QIAquick spin columns in a microcentrifuge. Even if distinct bands of the expected size are observed, primer dimers should be removed by PCR purification method. In the absence of the purification, the proofreading activity of DNA Polymerase will degrade the PCR fragments or remove the 3' terminal deoxyadenosine from the vector during ligation Reaction.

What is PCR?Polymerase Chain Reaction (PCR) is a powerful technique that has rapidly become one of the most widely used techniques in molecular biology because it is quick, inexpensive and simple.What is PCR used for?The technique amplifies specific DNA fragments from minute quantities of source DNA material, even when that source DNA is of relatively poor quality.How does it work?PCR can work with following components:An adequate amount of template DNA between 0.1 to 1 mg genomic DNA is required. It is worth to note that larger template DNA amounts usually increase the yield of non-specific PCR products.

Background: Pseudomonas lipopeptides such as white line-inducing principle (WLIP) have important antimicrobial activities. When a WLIP producer bacterium is grown close to P. tolaasii, a white line precipitate will be formed between them Identification of white line Reaction (WLR) defense strategy and its distribution among different pseudomonads might help to use such an approach for the treatment of infectious diseases In current study, through a degenerate Polymerase Chain Reaction (PCR) method, the partial characterization of genetic system of NRPS-based WLR was attempted in Pseudomonas aeruginosa for the first time in the world.Methods: Domain analysis of non-ribosomal peptide synthetase (NRPS) enzymes was performed by the nonribosomal peptide synthetase-Polyketide synthases (NRPS-PKS) tool. Multiple DNA sequence alignments, phylogenetic analyses and local Blast searches were performed by Geneious Pro. The gDNA was extracted from P. aeruginosa LMG 1272 and degenerate PCR was carried out.Findings: First, PCR amplification based on the C1 and TE domains was performed. Despite trying several modifications, a number of bands were obtained in addition to our expected band. For this purpose, the gene wlp/wip blast against Pseudomonas aeruginosa genomes was performed. Two homologes with about 50% identity to wlpB in RW10S2 were obtained. New primers were designed by which the target fragment could be amplified whose Blastp identified only one protein in bacterium Ralstonia solanacearum SD54 with the identity of about 43%.Conclusion: This study shows that the defensive white line formation by P. aeruginosa is governed most likely by a genetic system different from what has been reported before.

Background: Genotyping study and genetic diversity increase basic information about the parasites that cause malaria disease. These studies may support new treatment for disease, and control program in the future. Therefore, the present study was conducted to investigate the genetic structure and genotypes frequency based on MSP-3a gene of Plasmodium vivax in patients with Malaria reported in Isfahan City, Iran. Methods: Forty samples of patients infected with Plasmodium vivax, who were referred to laboratories of Isfahan health centers, were collected. The samples were tested using specific primers based on MSP-3a gene by nested Polymerase Chain Reaction (PCR) method. Findings: Based on PCR product size, three genotypes of PvMSP-3a gene as A (about 1900 bp), B (about 1600 bp), and C (about 1200 bp) were observed. The most frequent genotype was genotype A with 72%, and 10% of the patients showed both genotype A and B. Conclusion: The results show that PvMSP-3α genotype can be used as a genetic marker in both endemic and imported malaria areas. This gene also is acceptable as epidemiologic marker for diagnosis of Plasmodium vivax genotyping, and detection of mixed infection having both genotypes