Showing papers in "International Journal of Molecular and Clinical Microbiology in 2013"

Evaluation of biofilm-forming capabilities of urinary Escherichia coli isolates in microtiter plate using two different culture media

Abstract: The ability of uropathogenic E. coli (UPEC) to cause symptomatic UTI is associated with the expression of a variety of virulence factors. Biofilm formation enables UPEC to resist the flow of urine and increases its tolerance to antimicrobials and the host immune response. The measurement of biofilm formation in vitro is affected by the type of culture medium used. The aim of this study was to evaluate biofilm-formation capabilities of UPEC in microtiter plate using two different culture media. A total of 170 isolates of E. coli were isolated from patients with symptomatic UTI in Gorgan, north of Iran. Biofilm formation of the strains was examined in LB and BHI broth with the addition of 1% sucrose. The quantitative analysis of biofilm formation was performed using crystal violet staining followed by spectrophotometry measurement after addition of decoloring solution. The biofilm formation of UPEC isolates in LB broth (20 isolates; 11.8%) was significantly (p <0.001) lower than those grown in BHI broth (105 isolates; 61.8%). All isolates that formed biofilm in LB broth also formed biofilm in BHI broth. Whilst 36 (21.2%) isolates grown in BHI broth formed strong biofilm, only one (0.6%) isolate grown in LB broth exhibited a similar result (P<0.007). Our data suggest that the process of biofilm formation by UPEC is strongly modulated by culture conditions and the method employed. In our study the use of BHI broth supplemented with 1% sucrose proved to be superior to the LB broth and can be employed for measurement of biofilm formation in UPEC.

Antibacterial activity of the sea cucumber holothuria leucospilota

Abstract: Article history: Aquatics are a source of bioactive compounds that these compounds have different properties such as antimicrobial activity. In this study, antibacterial activity of methanol, chloroform and hexane extracts from body wall, gonad and intestine of sea cucumber Holothuria leucospilota was evaluated against Bacillus subtilis, Pseudomonas aeruginosa and Staphylococcus aureus. Bioactive compounds of body wall, gonad and intestine of the sea cucumber Holothuria leucospilota collected from the north coast of the Persian Gulf were extracted using methanol, chloroform and hexane. The antibacterial activity was determined using the serial dilution method. The minimum inhibitory concentration and minimum bactericidal concentration was evaluated by broth micro dilution method. Results demonstrated that the P.aeruginosa was shown to be the most sensitive microorganism. All concentrations of methanol extracts from body wall, gonad and intestine did not show antibacterial activity against B.subtilis and S.aureus. Methanol extracts of gonad and intestine and chloroform extract from body wall showed no antibacterial activity against P. aeruginosa. Hexane extract from gonad had no inhibitory effect on the growth of B. subtilis in any of the concentrations. Other extracts had antibacterial effect in certain concentrations studied. None of the extracts showed any bactericidal effect against B. subtilis. Based on findings of this study, sea cucumber extracts can be considered as a natural antibiotic in the future research.

A study on the frequency of vaginal species of Mycoplasma genitalium, Gardnerella vaginalis and Neisseria gonorrhoeae among pregnant women by PCR technique

Abstract: Article history: Gardnerella vaginalis, Mycoplasma genitalium, Neisseria gonorrhoeae, bacterial vaginosis, Polymerase Chain Reaction (PCR), Bacterial vaginosis or non-specific vaginitis describes the disease caused by a change in the normal Flora of the vagina, which leads to the elimination of Lactobacilli, generating hydrogen peroxide and excess growth of bacteria, particularly anaerobic bacteria. This disease is the most prevalent infection of the female genital tract, and the rate of frequency of anaerobic bacteria, specifically vaginal species of Gardnerella and Mycoplasma, is 100 to 1,000 times higher than that of healthy individuals. To determine the rate of frequency of Gardnerella vaginalis, Mycoplasma genitalium and Neisseria gonorrhoeae, which are present in bacterial vaginosis. samples of vaginal secretions of pregnant women referred to the Women's Clinic in the Tonekabon Township were obtained. In order to detect the presence of Gardnerella vaginalis, Mycoplasma genitalium and Neisseria gonorrhoeae, the samples were studied using the Polymerase Chain Reaction (PCR) method. After obtaining the data, the results were analysed using the Chi-square (x2) test. Of the 44 samples tested, 3 cases were found to contain Gardnerella vaginalis (6.81 percent), 2 cases to contain Neisseria gonorrhoeae (4.54 percent), and 10 cases to contain Mycoplasma genitalium (22.72 percent). Statistical analysis showed that Mycoplasma genitalium was significantly related to the consequence of abortion. However, there was no relationship between infections caused by Gardernerella vaginalis, Mycoplasma genitalium and Neisseria gonorrhoeae with premature delivery and hospitalization of the newborn in the Neonatal Intensive Care Unit (NICU). Considering the findings, it seems that a low percentage of the studied populations were afflicted by the bacterial vaginosis.

Serological survey of leishmania infantum in wild rodents of turkemen sahra, golestan province, north-east iran

Abstract: Leishmania infantum, an etiologic agent of zoonotic visceral leishmaniasis (VL), is spreading in Iran. Dogs are the main reservoirs but L.infantum has been detected in jackals, foxes and wild rodents too. Despite high number of rodents in the world, the exact role of them in epidemiology of VL is not clear. In recent years the number of sporadic cases of dogs and humans VL has been increased in arid and semi-arid areas of Golestan province. The aim of this study was to detect L.infantum antibodyin wild rodents of Turkemen Sahra district of Golestan province. 75 rodents were randomly trapped from 8 areas and after identification of the genus and species, blood was collected by cardiac puncture. Indirect immunofluorescence antibody test (IFAT) was performed on serum samples of captured rodents. % 8 of serum samples which belonged to Mus musculus population were positive. Results of current study highlight the possible role of wild rodents in the spreading of L.infantum in ecosystem. More epidemiological and molecular study is recommended to imagine the exact role of wild rodents in life cycle of L.infuntum in Golestan province.

Propolis, royal jelly and pollen from beehive have antibacterial effect on aquatic pathogenic bacterial isolates

Abstract: Article history: New approaches for treatment of infectious diseases in aquatic animals have important roles in aquaculture technology progress. In the present study, In vitro effects of different extracts of propolis, royal jelly and pollen obtained from beehives have been investigated on aquatic pathogenic bacterial isolates. The isolated bacteria identified on the basis of their biochemical properties and sequence alignment of the amplified genome fragments. Antimicrobial activities of ethanol extracts of royal jelly, propolis and pollen, and acetone extract of propolis were determined through well diffusion and microdilution methods. The isolated bacteria identified as Aeromonas and Vibrio spp., based on biochemical characterization. Alignments of the amplified sequences showed most similarites to Vibrio cholerae and Aeromonas hydrophila. The results obtained from antibacterial effects of extracts showed that the acetone extract of propolis as well as the ethanol extract of royal jelly, had the greatest effect on Aeromonas hydrophila (MIC=25 mg ml -1 ); and the ethanol extracts of pollen and royal jelly as well as the acetone extract of propolis, showed the greatest effect on Vibrio cholerae (MIC=50 mg ml -1 ). The results of present in vitro study propose the beehive compounds (royal jelly, propolis and pollen) as powerful natural products to control pathogenic bacteria in aquaculture systems.

Molecular epidemiology of viral meningitis in children in south east of Caspian Sea, Iran

Abstract: Viruses are the most common causes of aseptic meningitis. Early detection, treatment and management of viral meningitis are priority. This study aimed to evaluate common viral meningitis in children referred to Taleghani pediatrics hospital in Gorgan, south east of Caspian Sea, Iran. In this descriptive study CSF and blood samples were taken from 40 children with negative bacterial culture who were referred with meningitis symptoms since Jun 2008 till Sep 2010. Samples were used for viral, biochemical and cytological assays. DNA extraction was done by high pure viral nucleic acid kit of viral nucleic acid from CSF. PCR and Real-time PCR were performed for detection of viruses. Demographic, clinical, biochemical and cytological data were collected and entered in SPSS version 18. All cases with p<0.05 were considered as significant. In overall 12 (30%) viruses were detected by distribution of 5 (41/7%) Enterovirus, 4 (33/3%) Herpes simplex virus-1 (HSV-1) and 3 (25%) Mumps virus. Patients aged between 1 month to 10 years old with mean of 3 years old of which 92/5% were living in urban area. All positive cases showed fever and CSF Pleosytosis with no bacterial growth, gram staining and urinary tract infection. In conclusion, the results showed that clinical and biochemical analyses are not sufficient for certain diagnosis of meningitis in children and molecular assay is recommended to apply for early detection, treatment and management of viral meningitis.

In vitro evaluation of nickel oxide-based nanocomposite as wound dressing material against the bacterium isolated from burns

Abstract: The introduction of newly devised wound dressing has been a major breakthrough in the management of wounds or infections. The aims of this paper are to isolate and identify bacterial species causing burn wound infections from a University-related Iranian hospital as well as determination of the antimicrobial susceptibility of the isolated microorganisms to newly devised nanocomposite materials for developing efficient wound dressing. The NiO nanoparticles were generated in situ and subsequently impregnated on the surface of cotton fabrics using ultrasound irradiation. Then, surface modification was performed to reduce initial bacterial attachment using polyethylene glycol. Cotton fabric was characterized by measuring scanning electron microscope (SEM), X-ray diffraction (XRD) and antibacterial properties. Disk diffusion method was used to quantify the efficacy of NiO-based wound dressing against the most common burn wound pathogen, Pseudomonas aeruginosa, isolated from burns and wound swabs patients of Emam Burn and Accidents hospital in Isfahan province, Iran. All isolates showed high resistance to the commonly used antibiotic (Ampicillin, Gentamicin, Cephalexin, Co-trimoxazole and Amoxicillin). In vitro evaluation showed that the modified cottons exhibited excellent biocidal action against high-resistant isolated Gram-negative bacteria compared to unmodified ones. The results suggested that NiO nanoparticles may be considered as an effective component of therapy for burn infections and in the combination with different antibacterial agents to overcome the resistance of the microorganisms and to obtain synergic antibacterial activity.

Frequency of Human papilloma virus among pregnant women by PCR technique

Abstract: Today, it has been confirmed that the viral infections play an important role in premature delivery and abortion. This study was carried out to determine the frequency rate of papilloma virus types 16 and 18 in pregnant women and their relationship with premature delivery and abortion. In this study, vaginal secretions were collected from pregnant women who had been referred to the women clinics in Tonekabon, a city in North of Iran. The samples were used to amplify E6 gene of papilloma virus through Polymerase Chain Reaction method. The results were analyzed by χ2 statistical test. Of 47 tested samples, 14 cases were infected with type 16 papilloma virus (HPV16) (29.78%), while 3 cases were infected with type 18 papilloma virus (HPV18) (6.38%). Papilloma virus infection had no significant relationship with abortion and premature delivery. Considering the obtained findings, it appears that a high percentage of the study population was infected with HPV16 virus. Therefore, risk of affliction with abortion and premature delivery can be decreased up to a limit in this region by screening before pregnancy.

Hwp1 gene Expression of Candida albicans and Study its role in adherence

Abstract: Candida albicans is an opportunistic fungal pathogen found as mycoflora in the human body surfaces. Sevral genes play a crucial rule in its virulence including Hwp1 (hyphal wall protein 1), BCR1 and ALS gene family. Hwp1 gene is a responsible for coding a cell surface protein, which mediates biofilm formation in candida albicans. Here we investigated the presence of the HWP1gene was characterised among Candida albicans isolates in women with recurrent vaginal canididasis and further we studies its role in cell adherence. We used 50 Candida albicans clinical isolates resistant to Fluconazole. RNA (of samples were extracted using glass bead and lysis buffer and cDNA was synthesized using reverse transcriptase enzyme. Expression of (HWP1) gene was analysed using reverse transcriptase-plolymerase chain reaction (RT-PCR). The ability of adherence of the isolates with or without the expression of HWP1 were characterized using Hela cells. Statistical analysis were performed using t-test and two-way ANOVA SPSS software. Our results showed that the HWP1 gene were present in 47 samples (94%) out of 50 isolates, 27 samples (57%) had expression of HWP1. The result of adherence assay revealed that the isolates with the expression of HWP1 gene and control isolates was statistically different (p<0.05). In conclusion, the isolates with the expression of HWP1 gene has the higher ability to adhere the epithelial mucosal cell surface. Our data support a positive correlation between the expression of HWP1 gene and the ability of adherence to epithelial cells.

Synthesis, Characterization and Antibacterial Activity Studies of Some N-Acyl-N'-aryl Thiourea Derivatives

Abstract: Article history: Received 14 May 2013 Accepted 21 November 2013 Available online 1 December 2013 Some N¬-acyl-N'-aryl thiourea derivatives 4(a-f) have been prepared by the reaction of acyl halides ammonium thiocyanate and aryl amines. The structures of synthesized compounds have been characterized by IR, 1HNMR spectral studies. The synthesized compounds 5(a-f) have been screened for antibacterial activity. The effect of the structure of the investigated compounds on the antibacterial activity is discussed.

Isolation and molecular identification of bacteria producing alkaline phosphatase enzyme from environmental sources

Abstract: Alkaline phosphatase (ortho phosphate, monoester hydrolase phosphoinositide E. C. 3.1.3.1) is a non-specific metalloproteinase enzyme that is located inside periplasmic space of bacteria. This enzyme is used to measure freshwater sediment in genetic engineering for cleaning water. Isolation and molecular identification of the bacteria producing alkaline phosphatase and comparison of its production rate in samples collected from different environmental sources including soil, wastewater and yogurt were the objectives of this study. Different environmental samples including soil, wastewater, stool and dairy products were cultured on the specific medium of phenolphthalein phosphate. Then, the bacteria producing the enzyme were isolated and identified based on the colony morphology, biochemical tests and finally PCT test and ribotyping. After the incubation, only the medium cultured with wastewater showed colonies that were discolored and pinkish. BLAST results of the samples confirmed the existence of Enterococcus durans, Shewanella putrefaciens and Shewanella xiamenensis; Shewanella putrefaciens with the highest concentration rate (87.48 u/l) was selected as the most superior strain. Investigating the enzyme concentration using a spectrophotometer at wavelength of 405 nm determined that the wastewater mediums were the only medium has potential to produce alkaline phosphatase enzyme among environmental sources of soil, wastewater, stool and yogurt. Other environmental samples did not show any potential for enzyme production.

Molecular Identification of Candida Species in Clinical Isolates

Abstract: Candida exists as a normal flora on skin and mucosa. The prevalence of Candida-induced infections specially mucosal and vaginal infections have been raised in the world in last decade due to increase in predisposing factors. C.albicans is the major etiological agent of candidiasis, but infections due to other species have been increasing. The aim of this study was to determine Candida species by using polymerase chain reaction (PCR) in patient with oral and vulvovaginal candidiasis that referred to clinics of Babol University of Medical Sciences, Babol, Iran. Sixty clinical samples were collected from mouth (no= 29) and vagina (no= 31). Identification of Candida species was done by using a combination of conventional and PCR methods. Germ tube and vesicle (chlamidoconidia) were seen in 43(71.67%) and 42(75%) of Candida isolates, respectively. Assimilation of sugars and Gel electrophoresis of PCR products showed that 78.34% and 75% of isolates were C.albicans, respectively, followed by C.parapsilosis, C.guilliermondii, C.tropicalis and C.glabrata.According to our results, C.albicans was the major causative agent in mouth, and also in vulvovaginal candidiasis. We concluded that Candida spp. is prevalent in mouth and vagina, indicating the necessity for preventing candidiasis by using some antifungal drugs.

Monitoring subtypes of the human polyomavirus BK in Iranian adult kidney transplant patients

Abstract: BK virus (BKV) is a polyomavirus with seroprevalence in adults, ranging from 60 to 100%. It is considered as usual cause of renal dysfunction after the allograft renal transplantation nephropathy. Potent immunosuppressive therapy in kidney transplantation can lower the rate of acute rejection. Therefore, untreated BKV infections lead to kidney allograft dysfunction or loss. In order to estimate the difference, this study investigated the BKV in urine samples of kidney transplant patients. In this study, we used 220 urine samples from allograft recipients in the time period of 2010-2013. Then, the 287 bp typing region and the PCR increased from the urinary DNA. The PCR products were digested by three limitation enzymes, namely AluI, Cfr13I and RsaI to determine the BKV subtypes. The BKV subtypes are common in the city of Esfahan, Iran. This research showed that 102 (75%) samples were infected by BKV type I. 7 (5%) and BKV subtypes II, 5 (4%) III, and 22 (16%) IV were found in our patients. On the other hand, mixed infections did not clear in the recipients. Our findings showed that BKV replication might occur after kidney transplantation and through the early hours. BKV types II, III and IV are brand new in Iran and previously were not apparent in samples of urine in different kidney transplant patients.

Survey of antibiotic susceptibility pattern in microorganisms isolated from clinical samples in shahid faghihi hospital of shiraz, iran

Abstract: 1Department of Microbiology, School of Public Health, Tehran University of Medical Sciences. Tehran, IR Iran. 2Department of Microbiology, Faculty of Medicine, Iran University of Medical Sciences. Tehran, IR Iran. 3– Department of Mycology, Faculty of Medicine, Mazandaran University of Medical Sciences. Sari, IR Iran. 4Department of Food Microbiology, School of Public Health, Tehran University of Medical Sciences. Tehran, IR Iran. 5Department of Microbiology, Faculty of Medicine, Iran University of Medical Sciences. Tehran, IR Iran.

Antagonistic activity of Streptomyces gresiofuscus PTCC1628 against isolated Gram negative bacteria from Urinary Tract Infections

Abstract: Article history: Received 27 May 2013 Accepted 7 Agust 2013 Available online 1 December 2013 Antibiotics are the best known products of actinomycete Over 5,000 antibiotics have been identified from the cultures of Gram positive and Gram-negative organisms, and filamentous fungi, but only about 100 antibiotics have commercially been used to treat human, animal and plant diseases The genus, Streptomycete, is responsible for the formation of more than 60% of known antibiotics Bacterial urinary tract infections are frequent in the outpatient as well as in the nosocomial setting The common bacteria from UTIs were isolated from hospital and laboratory samples The present study was designed to isolate Gram-negative bacteria from urinary tract infections and evaluate Streptomyces gresiofuscus PTCC1628 antimicrobial activity against pathogenic bacteria Bacterial isolates were identified as Acinetobater spp and Pseudomonas aeruginosa The antimicrobial activity was examined by the agar Well diffusion method The cell free supernatants of the Sgresiofuscus were able to inhibit the growth of all human pathogens (Acinetobacter spp and Paeruginosa) isolated in this study in Well diffusion method The isolates also showed very promising activities against multi drug resistant human pathogens Concentrations of produced compounds by Sgriseofuscus PTCC1628 were determined by GC-MS method

Response of surface optimization for the enhanced production of alkaline protease isolated from bacillus sp. with bean husk as a new substrate

Abstract: Article history: Received 2 June 2013 Accepted 17 November 2013 Available online 1 December 2013 Optimization of the fermentation medium for maximum alkaline protease production was carried out. Fifteen positive isolates were examined for their extent of alkaline protease production. The most potent producer was identified as Bacillus sp. The solid substrate screening showed that the combination of wheat straw and bean husk was the best one. The initial screening by using Plackett-Burman's design demonstrated that among the tested factors, casein, ammonium sulphate and pepton as the nitrogen sources and glucose, lactose and sucrose as carbon sources, glucose and casein significantly (P < 0.05) enhanced the protease production in combinatory solid state fermentation (SSF). Further optimization of protease production by Bacillus sp. strain k7 on different factors such as incubation time (84 h), inoculums size (64%), initial moisture content (97%), buffer volume (4.9%) in SSF by applying RSM was achieved.

Identification of cutaneous granuloma caused by Mycobacterium marinum using PCR method

Abstract: Article history: Atypical Mycobacterium granulomatous skin infections are often accured by Mycobacterium marinum, M. ulcerans, M. fortuitum, and M. avium colonies. Skin infections probably originate from an environmental source such as contacting with aquatic animals, fish farming and swimming in the pools, and inoculate into skin through skin wounds, scratches, trauma, and surgery. The lesions appear as purple papules, nodules in hands and feet, plaque blisters wart ulcers and markers transmission (sporotrichosis) in the path of lymph nodes. They have granulomatous accumulation with giant cells, and abscess pus appears, and sometimes in the form of ulcerative. Infection is limited to the skin, while in immunosuppressed cases it would be able to infect the whole body. To determine if Mycobacteria were present in granulomatose skin lesion, a total of 58 paraffine embedded tissue blocks were obtained and their DNA was extracted. The polymerase chain reaction (PCR) was used to amplify the HSP-65 gene. PCR amplification demonstrated the presence of Mycobacterium spp. In 18 blocks (31%). Among these 18 blocks, 8 (44%) were positive for M. marinum, 3 (17%) for M. ulcerans, 5 (27%) for M. fortuitum and M. chelonae, and 2 (12%) for M. avium. We conclude that Mycobacteria ought to be considered in the treatment of skin granulomas in Iran.

Survey on viability of streptococcus iniae in frozen rainbow trout (oncorhynchus mykiss) following experimental infection

Abstract: Streptococcus iniae is one of the most important agents that causes high mortality and losses in rainbow trout farms in Iran and the world. These bacteria can affects the consumers and people who deal with aquaculture via affected fish. So, viability of this bacterium in frozen fish and fish product can be useful for health care of consumers. A total of 90 rainbow trout, with average weight of 50 ± 3 g, were supplied from a fish farm and were transferred to six 200- liter tanks (15 fish per tank). The tanks were divided in two groups as control and treatment. The treatment group fish were experimentally challenged to Streptococcus iniae at dose of 3.6 × 105 cell/fish via intraperitoneal injection. Control group fish just received 0.1 ml of normal saline (0.9 % NaCl). In treatment group after 120 hours 91% of the fish were died and then mortality remained constant up to 14 days post challenge. Moribund and dead fish of treatment group and fish of control group were frozen and then kidney, liver and brain of fresh dead and frozen fish were used for bacterial culture after 8, 14, 20, 26, 30, 34, 36 and 38 months post freezing, to investigate the viability of these bacteria in different organs at freezing temperature (-20˚C). Streptococcus iniae were re-isolated from all of fresh and frozen tissue specimens within 38 month after freezing. All bacterium species confirmed as Streptococcus iniae using polymerase chain reaction (PCR) method. According to this study this bacterium as a zoonisis can be survived at least 38 month in freezing temperature (at -20˚C) in fish products, so the potential pathogenicity of this bacterium should be concerned in affected area as well as imported frozen fish.