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Background: Pili producing genes in different life cycles of Mycobacterium tuberculosis (M. tuberculosis) were assessed. M. tuberculosis has two life cycles: dormant and active states. We aimed to assess the pili producing genes such as curli pili of M. tuberculosis (mtp) encoded by the mtp gene (Rv3312A) and fimbrial low-molecular-weight protein encoded by flp gene (Rv3656c) which were compared and analyzed. Methods: Two hundred M. tuberculosis isolates were investigated both at active and dormant states for production and expression of pili. The dormant M. tuberculosis was achieved by incubation in a sealed tube (modified Wayne method). The susceptibility of M. tuberculosis was evaluated on genes, rpob, inh, katg, and gyra by using multiplex polymerase chain reaction (PCR) and single-strand conformational polymorphism methods. The PCR–restriction fragment length polymorphism was used to express pili genes mtp and flp and then the PCR products was digested using restriction enzyme Fnu4HI, XmaI, and MspJI and AciI, TagII, and HaeII, respectively. The transmission electron microscopy was also used to detect pili in different isolates. The result was compared and analyzed using H37RV as a gold standard. Results: The mtp and flp PCR products were 263 and 122 bp in the studied strains irrespective of M. tuberculosis different life cycles, respectively. The PCR products were analyzed on 8% Polyacrylamide gel electrophoresis (PAGE), and in the 180/200 (20%), producing five fragments of 25,40,45,63,90 bp with the Fun4HI and two fragments of 126,138 bp with the XmaI and uncut with the MspJI for mtp gen were obtained at the dormant and active states of M. tuberculosis (P < 0.05). Similarly in flp gene producing three fragments of 22,35,65 bp with AciI and two fragments of 35.87 bp with TagII and two fragments of 38.84 bp with HaeII were obtained (P < 0.05). In contrast to genotyping analysis, the electron microscopy examination showed protruding of pili from M. tuberculosis, especially in dormant mycobacterium (15/100; 15%), that was multidrug resistance and extensive drug resistance isolates (P > 0.05). Conclusion: Pili were shown by electron microscopy, although at the gene expression, the insignificant difference was observed at the dormant strains in comparison to active states. Therefore, we may conclude that other genes might be involved in pili production of M. tuberculosis that needs further investigation. Although, the resistance phenomena might influence the pili producing gene expression that showed in our results.

Objective(s): Anti-tumor effects of Lactobacilli as normal flora have been described. In a previous study, we identified a protein isolated from the bacterium Lactobacillus casei ATCC 39392 in acidic pH conditions named metallopeptidase. Therefore, we decided to evaluate the effect of the recombinant plasmid coding metallopeptidase protein on the inhibition, proliferation, or apoptosis of the colorectal and breast cancer cell lines. Materials and Methods: Identified metallopeptidase gene of L. casei under the specific colon cancer promoter was transferred to the Human SW480 and MDA-MB231 cells. Cell viability was evaluated in these two cancer cell lines via MTT assay, apoptotic changes, and expression level of p53 and MAP2K1 genes in comparison with healthy blood cells as a control group. Results: Viability of SW480 and MDA-MB231 cells was identified at 25% and 7%, respectively. An increase in apoptotic cell death in the SW480 cell line was observed as revealed by Tunnel staining. The expression assay of TP53 and MAP2K1 genes showed that MPL protein altered gene expression in a cell type-specific manner. Tunnel analyses showed that the pronounced cytotoxic effect of pEGFP-C2/MPL plasmid on SW480 cells was mediated through apoptosis. Conclusion: These results suggest that endogenous recombinant MPL under colon specific promoter inhibits the proliferation of SW480 colorectal cancer cells by increase in MAP2K1 and P53 activation. L. casei metallopeptidase under the same circumstances could not affect the growth rate and viability of MDA-MB231 breast cancer cells in vitro.

Stenotrophomonas maltophilia is resistant to a wide range of antibiotics. This study aimed to investigate the beta-lactamase, ESBL, and MBL enzymes and also L1and NDM1 genes in isolates of Stenotrophomonas maltophilia. Antibiotic susceptibility test, beta-lactamase, ESBL, and MBL test was done on 23 isolated Stenotrophomonas maltophilia. The presence of L1 and NDM1 genes was investigated on isolated MBL bacteria by the PCR method. The results showed the most effective antibiotics were tigecycline and gentamicin (100%) and the highest resistance was observed with aztreonam (56.22%). 39.13% of isolates were ESBL, 82.66% of isolates were beta-lactamase positive, 60.86% of isolates were Metallo-beta-lactamase positive. 39.13% of MBL positive isolates, were positive for the L1 gene but the NDM1 gene was not seen. Results of this study showed the higher resistance to beta-lactam antibiotics also the presence of the l1 gene. ESBL and MBL producing S. maltophilia are frequently resistant to a wide range of antibiotics. Therefore, needs to be done antibiotic sensitivity test, ESBL, and MBL test before treatment for this bacterium.

Mycobacterium tuberculosis has infected more than two billion individuals worldwide, of whom 5%–10% have clinically active disease and 90%–95% remain in the latent stage with a reservoir of viable bacteria in the macrophages for extended periods of time. The tubercle bacilli at this stage are usually called dormant, non-viable, and/or non-culturable microorganisms. The patients with latent bacilli will not have clinical pictures and are not infectious. The infections in about 2%–23% of the patients with latent status become reactivated for various reasons such as cancer, human immunodeficiency virus infection, diabetes, and/or aging. Many studies have examined the mechanisms involved in the latent state of Mycobacterium and showed that latency modified the expression of many genes. Therefore, several mechanisms will change in this bacterium. Hence, this study aimed to briefly examine the genes involved in the latent state as well as the changes that are caused by Mycobacterium tuberculosis. The study also evaluated the relationship between the functions of these genes.

Background and Objectives: In recent decades, enterococcal resistance to antimicrobials has greatly increased. Furthermore, these chemicals include several side effects on the patients. Since no reports are available of the bacteriophages' effects on eukaryotic cells, they can be good solutions for multidrug-resistant bacterial problems. Therefore, the major aim of this study was to isolate bacteriophages from wastewaters on clinical antibiotic-resistant enterococci. Materials and Methods: Clinical bacteria were isolated, then enterococcal isolates were identified using different methods. The antibiotic resistance scheme of the enterococcal isolates was assessed. The bacterial isolates were exposed to wastewater samples containing potential bacteriophages. Technically, isolated bacteriophages were studied by electron microscopy. Results: Isolated bacteria were verified as Enterococcus faecium. Results showed that bacteriophages could easily be isolated from wastewater sources. The isolated bacteriophages were effective on E. faecium as well as Streptococcus dysgalactiae. Furthermore, these bacteriophages were challenged with five other bacteria (ATCC) with no visible effects. In general, the isolated bacteriophages belonged to the Myoviridae, Siphoviridae, and Inoviridae families. Conclusion: Further studies on bacteriophages and their efficacy on enterococcal strains could increase the treatment possibility of enterococcal infections. Due to these bacteriophages' effects on Streptococcus strains, bacteriophages may be used to treat streptococcal infections as well.

Abstract Sarocladium kiliense is a soil saprophytic mold with worldwide distribution, which can infect humans and other mammals, sporadically. The clinical manifestations include mycetoma, onychomycosis, keratomycosis, pneumonia, and arthritis. Here, we present a disseminated infection due to S. kiliense in a diabetic patient infected to coronavirus disease 2019 (COVID‐19) from Isfahan, Iran.

Background and Objectives: The microbial contamination of wastewater is associated with health risks. The aim of this study was to use the autochthonous Bdellovibrio potential to prey Gram-negative pathogenic bacteria as a bio-control agent to treat urban wastewater. Materials and Methods: Thirty-six raw sewage samples were collected for isolation of Bdellovibrio. Double layer plaque assay was used for isolation and the isolates were identified by microscopic examination and molecular analysis. To evaluate the predatory potential for decrease number of Gram-negative pathogenic bacteria, plaque perdition assay, reduction in host cells viability by colony-forming unit (CFU) counting, reduction in optical density (OD) in co-cultures and assay of killing efficiency were carried out. Also, the raw wastewater was treated by Bdellovibrio then the reduction in CFU counting and reduction in OD was evaluated. Results: Four strains of Bdellovibrio were isolated and were registered in Gene Bank. Clear plaques were observed after 3-6 days of incubation for all prey cells. The CFU enumerations of all preys were decreased after 48 hrs in co-cultures and raw wastewater. Also, OD was decreased down to 0.2 nm after 48 hrs. Conclusion: These autochthonous Bdellovibrio strains are proposed to use for bio-control of Gram-negative pathogenic bacteria in wastewater and reuse it for irrigation in arid regions.

Background and Objectives: Lactobacillus casei, an acid-resistant bacterium, has a protective role against the pathogens. So we aimed to determine the proteome of Lactobacillus casei ATCC39392 strain in response to different pHs of 5 and 7 using proteomic analysis. Materials and Methods: Supernatant and bacterial extraction of Lactobacillus casei ATCC39392 adapts at pHs 5 and 7 were isolated using sodium dodecyl sulfate–polyacrylamide gel and two-dimensional gel electrophoresis. The comparison of results showed that 7 protein spots were seen in pH 5 but not in pH 7. Afterward, they were excised and sent for Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) to be identified. Results: Seven different proteins (four secretory and three structural) with different roles in human body health were identified. Prescribed proteins include putative cell wall associated Hydrolase, Glycoside Hydrolase, beta-N-Acetyl hexosaminidase, Histidine Kinase, Chaperonin, metal dependent Hydrolase and Lysozyme. Conclusion: Seven isolated proteins with anti-cancer and digestive impresses are proper subjects in therapy or drug delivery approaches especially oral drug usage for protection against stomach acidic area.

Objective(s): Pyocyanin, a blue-green pigment produced by Pseudomonas aeruginosa, interferes with host redox cycles, which can lead to generation of reactive oxygen species and progressive cellular oxidative damage. The aim of this study was to assess the influence of pyocyanin on human pancreatic cancer cell line.Materials and Methods: Polymerase Chain Reaction (PCR) was applied to confirm the existence of a specific pyocyanin producing gene (phzM). The pigment was then characterized by UV-visible, FT-IR, and HPLC analysis. Panc-1 cells were treated by different concentrations of pyocyanin and their cytotoxic effect as well as the induction of apoptosis/necrosis were evaluated by XTT assay and flow cytometry.Results: An overnight pyocyanin treatment resulted in significant cell reduction in a concentration-dependent manner. Inhibition rate of 6 mg.ml-1 pyocyanin (the highest concentration) extracted from clinical and soil isolates of P. aeruginosa were 98.69±0.23 and 89.88±1.86%, respectively, which decreased as the pyocyanin concentration lessened. Pyocyanin could also induce dose-dependent apoptosis/necrosis in Panc-1 cells after 24 hr.Conclusion: We reported, for the first time, cytotoxic effects of pyocyanin against human pancreatic cancer cell line. Considering this effect of the pigment, study on pyocyanin as a potential anti-tumor biodrug requires further studies.

Objective(s): Pyocyanin is a blue-greenish redox-active pigment, produced by Pseudomonas aeruginosa, with a wide range of biological and biotechnological applications. Pyocyanin biosynthesis is regulated by the quorum-sensing (QS) system in which the expression of QS genes and QS-controlled virulence genes may be affected by serum as a complex medium. In the current study, effects of adult bovine serum (ABS) and fetal bovine serum (FBS) on the production of pyocyanin were examined in order to develop it. Materials and Methods: The presence of pyocyanin-producing specific genes and proteins in clinical and soil isolates of P. aeruginosa was confirmed using PCR and SDS-PAGE. Isolates were inoculated to media containing different concentrations of complement-active/-inactivated ABS or FBS and pyocyanin concentration was measured by spectrophotometry. Extracted pigment was characterized by using UV-Visible spectrophotometry. Titration of ABS antibodies against studied isolates was performed by the tube agglutination test. Results: Adding ABS to P. aeruginosa culture medium decreased pyocyanin production compared to the control, while its production increased in FBS-containing media (113.21±2.581 vs. 55.26±0.827 μg.ml-1 and 126.80±2.036 vs. 30.56±0.382 μg.ml-1 of C11 and E8 pyocyanin concentration in the presence of 10% FBS vs. control, respectively). Conclusion: In this study, due to the presence of inhibitors such as complement proteins and antibodies in ABS samples, the use of FBS devoid of antibodies was effective to increase pyocyanin production in studied isolates.

Objectives: To investigate possible sources of Stenotrophomonas maltophilia (S. maltophilia) in the clinical environment. Methods: Different samples were collected from Amol City of Iran. Steps for the identification of S. maltophilia included culturing, biochemical tests, polymerase chain reaction (PCR) of 16S rRNA gene and 23S rRNA gene. In addition, production of melanin pigment and patterns of motility of the bacteria, were also investigated. Results: In our study, 20 S. maltophilia strains were isolated from clinical sources, oxygen manometer apparatus of hospitals were 7/110 (6.36%), blood was 1/777 (0.13%), sputum was 4/40 (4%), urine was 1/2 947 (0.03%), tap water was 1/240 (0.42%) and dental suction was 6/120 (5%). The isolated bacteria showed production of melanin pigment with rates of strong, moderate, weak, and lack of pigment. Types of motilities were seen in isolates. Conclusions: The highest percentage of bacteria is isolated of oxygen manometer system and dental suction, yet has not been reported from oxygen manometer system. These bacteria have also been associated with patients who have respiratory problems, so it is essential for staffs of hospitals to draw attention to this source of bacteria.

Background: Staphylococcus aureus is considered as a major cause of skin and soft tissue infections, arthritis, osteomyelitis, infective endocarditis, and pneumoniae though community or nosocomial transmission. In this study, attempts were made to investigate the distribution of some important exotoxin genes, including hla, hlb, tsst-1, eta, etb, and etd among methicillin-resistant S. aureus (MRSA) isolated from a hospital patients in Tabriz, Iran. Methods: In the present cross-sectional study, a total of 90 S. aureus were isolated from children who admitted to a hospital during six-month in 2017. Isolates were identified using biochemical tests and then, using PCR, the isolates were tested for the presence of, hla, hlb, tsst-1, eta, etb, and etd genes. Results: It was found that 40% of the S. aureus were considered as MRSA strains by biochemical and molecular tests. The results of molecular detection of virulence determinants showed that eta, hla, etb, tsst-1, hlb and etd were detected in 86.1%, 80.5%, 30.5%, 27.7%, 22.2%, and 19.4% of isolates, respectively. Conclusion: Our findings clarify characterization of toxin production status of S. aureus isolates from patients in Iran. The current study showed that a majority of S. aureus isolates harbored eta and hla virulence gene.

The increasing prevalence of multidrug-resistant Klebsiella pneumoniae strains isolated from hospitals shows the limitation of recent antibiotics used for bacterial eradication. In this study, 81 K. pneumoniae isolates were collected from three hospitals in Tehran. Antibiotic susceptibility test showed the highest rates of resistance to cefotaxim (85.5%) and ceftazidime (78.3%), and the lowest rates of resistance were detected for colistin (16.9%), streptomycin (16.8%), and chloroamphenicol (21.7%). Eleven different resistance patterns were observed. Sixty-six out of 81 isolates (81.5%) were found to be multidrug resistant (MDR), and 35.8% of them belonged to A3 resistance pattern. 7.4% and 66.7% were KPC enzyme and armA gene positive, respectively. RAPD PCR assay of these bacteria showed 5 clusters, 16 single types, and 14 common types, and there was not any correlation between genetic patterns of the isolates and presence of resistance agents. Simultaneous detection of resistance-creating agents could be an important challenge for combination therapy of MDR K. pneumoniae-caused infections.

Backgrounds: Uropathogenic Escherichia coli strains are the predominant causative organisms of urinary tract infections (UTIs). Aminoglycosides are clinically useful antibiotics with bactericidal activity against this bacterium. The most common mechanism for resistance to these antibiotics are mediated through production of aminoglycoside modifying enzymes (AMEs). The most common of these enzymes are Aminoglycoside Acetyltransferases (AACs). The epidemiology of the dominant type of these enzymes, AAC(3)-II, varies from region to region. The aim of this study was to determine the antimicrobial susceptibility pattern with a focus on aminoglycosides and the prevalence of aac(3)-IIa gene among clinical isolates of uropathogenic Escherichia coli obtained from Delfan, Lorestan, Iran. Materials and Methods: In this descriptive study, a total of 100 uropathogenic Escherichia coli isolates were collected from BoAli hospital in Delfan city, Lorestan, from July to November 2010. Antibiotic susceptibility patterns of the isolates were determined using disk diffusion method according to Clinical and Laboratory Standards Institute )CLSI) guidelines. Prevalence of aac(3)-IIa gene was determined by PCR and the relationship between resistance phenotypes to aminoglycosides and presence of aac(3)-IIa gene was evaluated. Results: Among the 100 tested isolates, maximal resistance was seen to ampicillin (85%); whereas, no resistance to imipenem was found. Sixty percent of the isolates demonstrated resistance to at least one of the tested aminoglycosides. Resistance rate towards these agents were as followed: gentamicin 39%, kanamycin 26%, neomycin 31% and amikacin 1%. Forty–four isolates (44%) harbored the aac(3)-IIa gene. The maximal rate of gene presence (36 isolates, 92.3%) was detected in strains with gentamicin resistant phenotype (39 isolates, 39%). Conclusion: On the basis of our findings, use of antibiotics such as nitrofurantoin, amikacin or imipenem are recommended for empirical treatment of UTIs. In addition, locally widespread distribution of aac(3)-IIa gene will pose concerns about progressive resistance against potent aminoglycosides such as gentamicin, tobramycin and others in the future.

Objective: Pyocyanine plays an important role in the pathogenesis of Pseudomonas aeruginosa, (P. aeruginosa) and is known to have inhibitory and bactericidal effects. This study has aimed to detect the phenazine biosynthetic operon (phz ABCDEFG) and two phenazine modifying genes (phzM and phzS) by polymerase chain reaction (PCR) and detection of its possible protein bands by sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE). The antimicrobial effects of pyocyanine alone and mixed with colloidal silver nanoparticles were studied.Materials and Methods: In this descriptive study, clinical and environmental species of P. aeruginosa were isolated by thioglycollate medium culture and cetrimide agar, respectively. The existence of a phenazine biosynthetic operon and two phenazine modifying genes as well as their protein products were confirmed by PCR and SDS-PAGE, respectively. Pyocyanine was extracted with chloroform and its antimicrobial effects against bacteria such as; Escherichia coli (E. coli), P. aeruginosaand Staphylococcus aureus (S. aureus) bacteria and yeast Candida albicans (C. albicans) were tested using well, spot and disk diffusion methods.Results: In this study, 3 out of 48 clinical strains were unable to produce pyocyanine on cetrimide and Mueller Hinton (MH) agar. Two strains did not have phenazine modifying gene bands. Another strain did not have the possible protein band of the phzM gene. Pyocyanine had antimicrobial effects against the microbial strains, which increased in the presence of silver nanoparticles.Conclusion: According to the results of the present study, some P. aeruginosa strains are unable to produce pyocyanine due to the absence of the phzM or phzS genes. Therefore, these genes have an important role in pyocyanine production in P. aeruginosa. Pyocyanine shows synergistic antimicrobial effects in the presence of silver nanoparticles against microbial strains.

Objective: The objective of this research was to determine the prevalenceof genital C. trachomatis infection in asymptomatic women by using highlysensitive nested-polymerase chain reaction (PCR) in urine sample.Materials and Methods: One hundred-forty asymptomatic women wererandomly selected from those who attended gynecology out patient departmentof Hazraat e Rasool Hospital in Tehran. First catch urine specimen were collectedfrom all the participants. DNA extraction was performed by means of HighPure PCR Template Preparation Kit (HPPTP) according to the manufacture’sinstructions. Extracted DNA was tested by omp1 gene based nested-PCR,using sets of primers to amplify C. trachomatis omp1 gene. Visualizationof a 1027 bp fragment from omp1 gene in agarose gel electrophoresis wasconsidered as a positive result.Results: In total, 140 urines were tested for determination of C. trachomatisinfection. C. trachomatis omp-1 was detected in 22.1% of cases (31/140). Theoverall prevalence rates of C. trachomatis in the urine sample as determined byomp1 based nested-PCR were 4.3% in group I (age, 44 years). The highest prevalence of C. trachomatis infection (12.1%) wasseen in women aged 25-34 years. This finding was not statistically significant(p=0.710). Also, there was not relation between C. trachomatis infection andsome probable risk factors such as young age (

Introduction: Shigella is a facultative intracellular pathogen that uses the host actin cytoskeleton protein for intra- and intercellular spread. The aim of this study was to determine the distribution of icsA gene and IcsA expressed protein bands among Shigella flexneri strains isolated from 3 clinical centers in Tehran. Material and Methods: Two hundred and seventy five isolated Shigella flexneri strains were identified by standard microbiological and biochemical methods. DNA isolation was performed using sodium perchlorate method. Hot start-PCR was done with 2 pairs of primers and the products were separated through agarose gel (0.8%) in TAE buffer. DNA fragments were visualized by ethidium bromide staining under UV illumination. Whole membrane preparation was used to examine the protein profiles and identification of probable IcsA (120-kda) protein band by SDS-PAGE. Results: From 100 isolated Shigella flexneri strains, both bands of 1600 bp and 1709 bp were detected in 46 isolates (46%). A 120 kDa band which seems to be related to IcsA protein was detected in 46 isolates (46%). The protein bands varied between 30 and 150 kDa. Discussion: IcsA is both necessary and sufficient for actin assembly in Shigella flexneri. Since icsA gene and IcsA protein band were not found in all Shigella strains, it seems that not all strains have the same pathogenesis. On the other hand, since the demonstration of icsA gene by PCR in all Shigella strains (46%) corresponded to the presence of a 120 kDa protein band by SDS-PAGE (46%), it seems that both tests may confirm each other. However, the PCR may be more accurate than SDS-PAGE.