Profiling of Virulence-associated Factors in Shigella Species Isolated from Acute Pediatric Diarrheal Samples in Tehran, Iran

OBJECTIVES: The genus Shigella comprises the most infectious and diarrheagenic bacteria causing severe diseases, mostly in children under five years of age. This study aimed to detect nine virulence genes (ipaBCD, VirA, sen, set1A, set1B, ial, ipaH, stx, and sat) in Shigella species (spp.) using multiplex polymerase chain reaction (MPCR) and to determine the relation of Shigella spp. from pediatric diarrheal samples with hospitalization and bloody diarrhea in Tehran, Iran. METHODS: Shigella spp. were isolated and identified using standard microbiological and serological methods. The virulence genes were detected using MPCR. RESULTS: Seventy-five Shigella spp. (40 S. sonnei, 33 S. flexneri, 1 S. dysenteriae, and 1 S. boydii) were isolated in this study. The prevalence of ial, sen, sat, set1A, and set1B was 74.7%, 45.4%, 28%, 24%, and 24%, respectively. All S. flexneri isolates, while no S. sonnei, S. dysenteriae, or S. boydii isolates, contained sat, set1A, and set1B. All isolates were positive for ipaH, ipaBCD, and virA, while one (1.4%) of the isolates contained stx. The highest prevalence of virulence determinants was found in S. flexneri serotype IIa. Nineteen (57.6%) of 33 S. flexneri isolates were positive for ipaBCD, ipaH, virA, ial, and sat. The sen determinants were found to be statistically significantly associated with hospitalization and bloody diarrhea (p = 0.001). CONCLUSION: This study revealed a high prevalence of enterotoxin genes in S. flexneri, especially in serotype 2a, and has presented relations between a few clinical features of shigellosis and numerous virulence determinants of clinical isolates of Shigella spp.

The Effect of Lactobacillus acidophilus PTCC 1643 on Cultured Intestinal Epithelial Cells Infected with Salmonella enterica serovar Enteritidis

OBJECTIVES: Gastrointestinal disorders caused by Salmonella enterica serovar Enteritidis (SesE) are a significant health problem around the globe. Probiotic bacteria have been shown to have positive effects on the immune responses. Lactobacillus acidophilus was examined for its capability to influence the innate immune response of HT29 intestinal epithelial cells towards SesE. The purpose of this work was to assess the effect of L. acidophilus PTCC 1643 on cultured intestinal epithelial cells infected with SesE. METHODS: HT29 cells were cultured in Roswell Park Memorial Institute medium supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. The cells were treated with L. acidophilus PTCC 1643 after or before challenge with SesE. At 2 and 4 hours post-infection, we measured changes in the expression levels of TLR2 and TLR4 via real-time polymerase chain reaction. RESULTS: Treatment with L. acidophilus inhibited SesE-induced increases in TLR2 and TLR4 expression in the infected HT29 cells. Moreover, the expression of TLR2 and TLR4 in cells that were pretreated with L. acidophilus and then infected with SesE was significantly higher than that in cells infected with SesE without pretreatment. Taken together, the results indicated that L. acidophilus had an anti-inflammatory effect and modulated the innate immune response to SesE by influencing TLR2 and TLR4 expression. CONCLUSION: Our findings suggested that L. acidophilus PTCC 1643 was able to suppress inflammation caused by SesE infection in HT29 cells and reduce TLR2 and TLR4 expression. Additional in vivo and in vitro studies are required to further elucidate the mechanisms underlying this anti-inflammatory effect.

Phage therapy: assessment of the efficacy of a bacteriophage isolated in the treatment of salmonellosis induced by Salmonella enteritidis in mice

Aim: This work aims to isolate and perform comparative studies of a phages active against a Salmonella enteritidis strain from Iran. Also, suitable phage candidates for therapy of mice will be selected

Background: Bacteriophage is of particular interest as a biocontrol agent in the prevention of food-borne illnesses. In recent years tend to use bacteriophages to control pathogenic bacteria has increased. A bacteriophage is considered to be a potent antibiotic alternative for treating bacterial infections

Methods: the specific phages against Salmonella Enteritidis was isolated and candidates for therapy of mice will be selected. Mouses divided into the six specific groups. Groups of mice were as follows: A: Bacteri [control] B: Bacteri+ bacteriophage [Simultaneous], C: Bacteri + bacteriophage Four days later, D: Bacteriophage + bacteri four days later E: Bacteri+ Ciprofloxacin [Simultaneous] F: Bacteri+ ciprofloxacin+ bacteriophage [Simultaneous]

Results: In this study, a lytic bacteriophage is isolated and it shows that phage has a head size of 46 nm and without a tail, by using an electron microscope. Oral administration of a single dose of 2 × 109 PFU/mouse bacteriophage enable to protect mouse against salmonellosis and it causes treatment of salmonellosis in mice

Conclusion: The use of this phage compared to ciprofloxacin shows that in addition of the treatment of mouse, it also prevents weight loss

Review of barrett's oesophagus to oesophageal adenocarcinoma: new prospect

Today's knowledge on oesophageal adenocarcinoma and its rising incidence has encouraged researcher to illustrate relationship between Barrett's disease and progression to adenocarcinoma. The incidence of this disease has been accelerated sharply in current decades since people life has changed. Studies have been demonstrated that several potential factors including genetical and environmental factors are involved on Barrett's transformation. Using different detectable biomarkers and techniques have supported early recognition of adenocarcinoma but still have not clarified pathogenesis of Barrett's oesophagus to oesophageal adenocarcinoma. This review summarizes as much as information in relationship with those evidences that have been finalised in different aspects of cellular and molecular pathogenesis of OA. However, current data shows that bacteria might play new role in the pathogenesis of adenocarcinoma

Isolation and identification of E. cowanii from powdered infant formula in NICU and determination of antimicrobial susceptibility of isolates

Enterobacter cowanii is a genus of common gram-negative, facultatively anaerobic, rod-shaped, non-spore-forming bacterium of the Enterobacteriaceae family. This organism can be potentially a powdered infant milk formula-borne opportunistic pathogen. The aim of this study was to isolate and identify E. cowanii from consumed powdered infant formula milk [PIF] in intensive care units [NICU] and to determine antimicrobial susceptibility patterns of this bacterium. E. cowanii was isolated according to FDA method in 125 samples of PIF milk purchased from drug stores between Jun 2011 and March 2012. For final confirmation, biochemical tests embedded in API-20E system were used. The drug susceptibility test was performed using the disc diffusion method according to CLSI recommendations. Out of the 125 PIF samples investigated, 4 [3.2%] samples were positive for E. cowanii. All four isolates from PIF samples were uniformly susceptible to imipenem, meropenem, ceftazidime, ciprofloxacin, and colistin. Fifty percent of isolates were resistant to ampicillin, amoxicillin, and cotrimoxazole. Analysis of the results indicated that complementary studies are necessary to clarify the possible role of E. cowanii as a food contaminant, in common NICU infections and high risk groups including persons with underlying disease and immunocompromised individuals

effect of reduced bacterial dilution on human amniotic membrane antibacterial activity, in vitro

The human amniotic membrane is the inner most layer of placenta and has antimicrobial effect, due to the presence of human beta-defensins and elafins. The purpose of this study is to investigate the effect of dilution reduction of 0.5 McFarland prepared from standard bacterial strains of Salmonella enterica BAA-708, Escherichia coli ATCC25922, Pseudomonas aeruginosa ATCC27853, Klebsiella pneumoniae ATCC7881, and Enterococcus faecalis ATCC29212 on antibacterial effect of human amniotic membranes in vitro. Amniotic membranes were obtained from the bank of organ transplantation in Imam Khomeini hospital, of women with elective cesarean section whose HIV, HBV, HCV and VDRL serological tests were negative. They were cut to 1.5x1.5 cm pieces. Then 0.5 McFarland suspensions of 1.5x10[8], 0.5x10[7] and 1.5x10[6] dilutions were prepared from bacteria which then were spread on Mueller Hinton medium agar and a piece of membrane was put in the center of each plate. After 24 hours incubation at 37[degree]C, the results were observed. In 0.5 McFarland standard dilution an inhibition zone was created in three standard strains of Pseudomonas aeruginosa, Escherichia coli, and Salmonella enterica unlike the other two strains. There was no change in the above results with two other dilutions and inhibition zone of sensitive strains was not created. Dilution reduction of microbial strains does not affect the antibacterial impact of amniotic membrane and dilution reduction does not yield to a false positive response and the conversion of resistant to sensitive strains

[Urinary tract infection in renal transplant patients in Sina university hospital]

Renal transplantation is the treatment of choice in patients with end-stage renal disease. Urinary tract infection [UTI] is one of the most common complications after renal transplantation and it has serious consequences. The aim of this study was assessing UTIs in renal transplanted patients and evaluation of risk factors associated with post-transplant UTI. In this prospective study, 173 patients [48 hospitalized patients and 125 outpatients] were enrolled in this study. These renal transplant recipients evaluated for bacterial urinary tract infection in urology research center at Sina Hospital. After collecting urine samples from symptomatic and asymptomatic patients, urinalysis and colony count were performed. Identification of bacteria was performed by routine microbiological tests in the Department of Pathobiology, School of Public Health, Tehran, Iran, in 2011. UTI was observed in 47 patients and the most prevalent microorganism was Escherichia coli [E.coli] 18 [38.2%]. Nearly 71% of UTI cases were diagnosed during the first three months post transplantation. Risk factors for post transplant UTI were female gender, age, length of hospitalization and diabetes mellitus. Female patients were more susceptible than males [OR=0.50 and P=0.047] to infection. There were no significant difference between diabetes mellitus and UTI. Most of the isolated bacteria were susceptible to imipenem and resistant to tetracycline and trimethoprim-sulfamethoxazole. Our study confirmed that bacterial infections remain as the most common infectious complication in the early post-transplant period, and antibiogram rather than empirical treatment is needed to find the best effective antibiotics. Moreover, risk factors such as female gender, increased age and length of hospitalization are predisposing factors to increased urinary tract infection in renal transplantation.

[Prevalence study of enterococus and staphylococci resistance to vancomycin isolated from urinary tract infections]

The role of gram-positive cocci especially Staphylococci species in causing urinary tract infection are well known. Among the Staphylococci species Methicillin Resistance Staphylococcus aureus [MRSA] is the most important. The rate of MRSA is increasing worldwide. This is alarming because the danger of these organism in public health. Therefore the aim of this study was to determine the sensitivity of gram-positive cocci, as well as MRSA to vancomycin and other antibiotics. This was a descriptive study, and were carried out on 300 patients with urinary tract infections[UTI] caused by gram-positive cocci, referred to Imam Khomeini hospital during eight months. Prior to the antibiotic sensitivity testing all the isolates were identified according to the standard conventional biochemical procedure, and then the antibiotic susceptibility test were carried out according to Bauer-Kirby method. Among the gram positive cocci causing UTI, the most abundant were Staphylococcus saprophyticus [37.7%], followed by Staphylococcus epidermidis [22.3%] and Staphylococcus aureus [18%] respectivley. The sex distribution of patients were 163 female [54.3%] and 137 male [45.7%] respectively, and the prevalence rate of urinary tract infections in female was[8.6%] higher than male. The rate of sensitivity of isolated Staphylococci were as followed, sensitive to vancomycine [100%], Ciprofloxacin [89.2%], rifampin [87.6%], and amikacin [71.8%] respectivley, but were resistant to penicillin and amoxicillin [100%]. The antibiotic sensitivity rate of isolated Streptococci was to vancomycine [85.1%], ciprofloxacin [50.7%] and penicillin [79.1%] respectively. Vancomycin is still a suitable antibiotic for the treatment of Staphylococcus infections. Although 6% rate of enterococci resistance to vancomycin is alarming, and use of this antibiotic in the treatment of other gram-positive bacteria should be done with precaution.

[ effect of amniotic membrane transplantation in healing of primary stages of pseudomonas keratitis]

Human amniotic membrane [HAM] forms the inner wall of the membranous sac that surrounds and protects the embryo during gestation. The main advantages of amniotic membrane transplantation [AMT] in the treatment of bacterial keratitis are its epithelial bandage properties. Previous studies have documented the presence of some antimicrobial proteins and peptides in amniotic fluid such as lactoferrin, lysozyme, bactericidal or permeability increasing protein, calprotectin [MRP8/14 protein complex], LL37, and neutrophil defensins [Human Neutrophil Peptides, HNP 1-3]. Furthermore, the amniotic membrane does not express HLA-A, B, C or DR surface antigens, which may help avoid rejection after its transplantation. Thus, it can be used as a biological immune barrier. The purpose of this study was to evaluate the effectiveness of the amniotic membrane's healing properties in rabbits with pseudomonas keratitis. By using an animal model, 14 rabbits were divided into two groups of controls and cases. A syringe was used to inoculate the corneal stroma of the animals by Pseudomonas aeruginosa ATCC27853. After 20 hours pseudomonas keratitis was created and amniotic membrane was transplanted to the cornea of the case group. The infiltration size were observed on the first, third and seventh days after the experiment. Corneal perforation was seen in the controls [P<0.001] but amniotic membrane prevented perforation in the case group [P=0.02]. Transplantation of amniotic membrane in the primary stages of pseudomonas keratitis treatment remarkably prevents corneal perforation and it can be used to control the disease process

Effect of amniotic membrane combined with ciprofloxacin in curing the primary stages of pseudomonal keratitis

Keratitis caused by Pseudomonas aeruginosa is often resulted in severe corneal ulcers and perforation, which leads to losses of vision. Human amniotic membrane [HAM] forms the inner wall of the membranous sac which surrounds and protects the embryo during gestation. The purpose of this study was to evaluate the effectiveness of the amniotic membrane's healing in rabbits with pseudomonas keratitis. In total 14 rabbits divided in 2 groups of: 1 as Control and 2 as experimental amniotic membrane combined with ciprofloxacin. A 0.05 ml suspension of Pseudomonas aeruginosa ATCC 27853 was injected into rabbit's corneal stroma, with no interference in control group. In the second group, the amniotic membrane in pieces of 1.5 x 1.5 cm transplanted to the entire corneal surface by eight interrupted 10.0 nylon sutures. In the first day ciprofloxacin drop was injected to the second group every 30 minutes and through second to seventh days every 2 hours. The results of perforation in cornea and the amount of infiltration were registered. The results showed that amniotic membrane transplantation [AMT] + ciprofloxacin group had 0% perforation and the control group 85.6%. Average infiltrations were 5 mm in AMT + ciprofloxacin groups and 23.75 mm in control. The use of amniotic membrane with ciprofloxacin was effective in prevention of cornea perforation and controlling the process of pseudomonal keratitis remission. The improvement of inflammation rapidly happened in ciprofloxacin + AMT group.

[Effects of probiotic Lactobacillus acidophilus and Lactobacillus casei on the behavior of colorectal tumor cells]

Probiotic microorganisms are living normal flora of human body that have nutritional value and health benefits when administered in adequate amounts. The health benefits include prevention of bacterial diarrhea, skin eczema and recently understood, prevention and control of various cancers, as well. Different mechanisms such as stimulating the immune system, modifying the composition of gastrointestinal and genitourinary tract normal flora and prevention of the carcinogenic activity of fecal enzymes have been identified for their probiotic activity. Due to the high density of the normal flora in the gut and also preferentially sporadic nature of colorectal cancers, these cancers are among the main candidates of treatment trials with probiotics. In this study, direct effects of probiotic lactobacilli on colon cancer tumor cells were studied. Supernatant fluid and bacterial extracts were prepared and CaCo-2 cells were treated by these materials. Subsequently, the effects of the aforesaid elements were evaluated on cell proliferation, cell necrosis and cell apoptosis by MTT assay, LDH assay and caspase-3 activity. The supernatants of lactobacilli decreased cell proliferation and increased cell apoptosis but they did not have any effect on cell necrosis. In contrast, when cancerous cells were treated by lactobacilli extract, it lead to cell necrosis in addition to reduction in cell proliferation and increase in cell apoptosis. The use of lactobacillus probiotics may reduce proliferation of tumor cells in the early stages of colorectal cancers

[ prevalence of extended-spectrum beta-lactamases and CTX-M-1 producing escherichia coli in urine samples collected at Tabriz city hospitals]

Numerous use of Beta Lactame in treatment of bacterial infections resulted in increments of drug resistance of such bacteria. One of difficulties in treatment of hospital infections is Extended Spectrum Beta Lactamase [ESBL] among isolated clinical strains of E.coli. Since some of ESBL strains shows double reaction in drug sensitivity test at in vitro and in vivo condition, therefore it makes difficulties in selection of right treatment. In the last years, CTX-M enzymes have become the most prevalent ESBLs in worldwide. The prevalence of ESBL types largely remains unknown in many parts of the Iran. This study was made to find the prevalence of ESBLproducing E.coli and molecular detection of CTX-M-1 in Tabriz. In the present study, 400 urine samples collected between November 2009 and April 2010, from Tabriz Hospitals were studied. Out of the 400 samples, 188 E.coli isolates were detected by standard biochemical tests. Susceptibility to antimicrobial agents was tested to 10 antibiotics by the disk agar diffusion [DAD] method. ESBL production was screened by phenotypic test that included both separate and combined disk agar diffusion techniques. The screened isolates were investigated by PCR assay to detect CTX-M-1 gene. From the total 188 E.coli isolates, 82 [43.6%] were shown to produce ESBLs by phenotypic test. During the PCR method on the 82 isolates, 69 [84.1%] were confirmed as CTX-M-1 producing group. The present study showed that CTX-M-producing isolates were increasing among E.coli strains and indicated the need for adequate susceptibility tests in laboratories for choosing the appropriate antibiotics for treatment

[Determination of genotype variation of Escherichia coli in well water of Tehran's parks by multiplex PCR]

Background and Aim: Water plays a major role in public health and the contaminated water has direct effect on human health. E.coli is an indicator of water pollution; different strains of E.coli such as ETEC, EHEC, EIEC and EPEC can cause diarrhea in children. The aim of this study is to investigate the contamination rate of well water of Tehran's parks, and pathotyping E.coli strains which cause diarrhea

Materials and Methods: 156 well water samples from five different geographical parts of Tehran [north, south, east, west, and the centre] were collected aseptically. The collected samples were transferred to the laboratory in microbiology section, School of Public Health, for membrane filtrations and patotyping

Results: 90 out of 156 samples [54/5%] were contaminated with E.coli. The samples from south part were more contaminated compared to others. Out of 90 isolated E.coli only 67 were DEC, in which 42 [62.7%] were detected as EPEC, 12 [17.9%] as STEC or ETEC, 9 [13.4%] as EIEC and 4 [6%] as ETEC by PCR method

Conclusion: Presence of different biotypes of E.coli in well water of parks can be dangerous, especially for children; thus, placing warning signs and educating people in this regard should be taken seriously

Diarrhea caused by enteropathogenic bacteria in children

Diarrhea is a major cause of mortality in 15 to 20% of the under 5-year-olds, and among the bacterial agents causing diarrhea, the five most important are Shigella, Salmonella, E. coli, Campylobacter and Yersinia. In a one-year study conducted in cooperation with a local health center in Islamshahr, South of Tehran, rectal swabs were collected from children less than 5 years of age who presented with diarrhea. Frequency evaluations of Yersinia and Campylobacter along with other intestinal bacterial pathogens were made. A total of 1600 samples were analyzed, 235 [14.7%] of which were positive. Most [119 cases; 50%] high-risk ages included children less than 2 years of age. Enteropathogenic Escherichia coli [EPEC] was the most common [109 cases; 6.8%] cause of diarrhea followed by Shigella [54 cases; 3.4%], Salmonella [46 cases; 2.9%], Campylobacter [15 cases; 0.9%] and Yersinia [11 cases; 0.7%]. Seasonal investigations show that, with the exception of Yersinia, all pathogenic bacteria are the most frequently isolated organisms occurring in the summer. Better knowledge about the causes of infection and the epidemiological situation could help the management of intestinal infections