Curcumin carbon dots inhibit biofilm formation and expression of esp and gelE genes of Enterococcusfaecium.

The increasing prevalence of vancomycin-resistant Enterococcus faecium, along with the ability of this bacterium to form biofilm on biotic surfaces and medical devices, has created a serious challenge. Therefore, the development of new antibacterial agents is an urgent need. In this study, curcumin carbon dots (Cur-CDs) were synthesized by a one-step hydrothermal method, and its antibacterial and antibiofilm effects were investigated. By broth microdilution method, the minimal inhibitory concentration (MIC) against vancomycin-resistant and sensitive clinical isolates of Enterococcus faecium (two clinical isolates in total) and standard strain of Enterococcus faecalis ATCC 29212 was determined, which were 1000, 1000, and 125 µg/ml, respectively. The inhibitory effect of Cur-CDs on biofilm formation of vancomycin-resistant E. faecium clinical isolates were evaluated by microtiter plate assay. Cur-CDs (1000 µg/ml) significantly prevented (p = 0.009) the biofilm formation of E. faecium isolates. Real-time PCR results showed that Cur-CDs (1000 µg/ml) significantly downregulated the expression of esp and gelE genes (p = 0.001 and p = 000000002, respectively) in clinical isolates of E. faecium, while Cur-CDs did not affect acm gene expression (p = 0.086). This study revealed that Cur-CDs can be effective antibacterial and antibiofilm agents against vancomycin-resistant and biofilm producer E. faecium, which makes them interesting candidates for treating or preventing bacterial infections.

Assessment of Helicobacter pylori positive infected patients according to Clarithromycin resistant 23S rRNA, rpl22 associated mutations and cyp2c19*1, *2, *3 genes pattern in the Early stage of Gastritis.

OBJECTIVE: Clarithromycin resistant Helicobacter pylori (CAM-R) is the main cause of standard triple therapy eradicating failure. Proton pump inhibitors (PPIs) directly pose bacteriocidic activity and prepare the optimum condition for Clarithromycin's best function. In counter with Poor metabolizer subjects, Homozygote Extensive Metabolizers have well characterized by treatment failure. Eventually, determination of CAM-R profile and estimation of PPIs metabolization rate support clinicians in better prescription. So, we explored Helicobacter pylori'mutations in 23S rRNA and rpl22 resistant genes, and cyp2c19 *1, *2, *3 allele variations, and PPIs metabolization patterns in patients, consequently the results reported to the physician. RESULTS: Sixteen out of 96 patients considered to be CAM-R Helicobacter pylori. A2143C (1/16), rpl22 insertion (16/16), and GTG deletion (2/16) recorded in CAM-R strains. P450 2C19 human genotyping demonstrated that the highest proportion of the H. pylori- positive strains infected patients 43/61(70.49%) categorized in Homozygote extensive metabolizer class. The rest (12/61)19.67% classified as Poor metabolizers, and 6/61(9.83%) distinct from Heterozygote extensive metabolizer group. Proportion of poor metabolizers and Heterozygote extensive metabolizer phenotypes between CAM-R strains mentioned to be 10/16(62.5%), and 6/16(37.5%). Cross points between the most frequently distributed allele in CAM-R strains indicated 81.25% for *2, and w2 for 18.75%.

Identification of Coxiella burnetii in Raw Milk of Livestock Animal in Iran.

Coxiella burnetii is the causative agent of Q fever in humans and animals. This study aimed to determine the frequency of C. burnetii in milk samples of dairy animals (goats, sheep, and cattle) in some selected regions in Iran, where there is no information about prevalence of C. burnetii. In this study, 162 individual milk samples were collected from 43 farms in three provinces (Tehran, Hamadan, and Mazandaran). Real-time PCR was used for the detection of IS1111a element of C. burnetii. In total, 23 of 162 samples (14.2%, 95% confidence interval (CI): 9.65-20.2%) were positive for C. burnetii by real-time PCR. C. burnetii was detected in 10.17% (95% CI: 4.74-20.46) of goat milk samples. In sheep milk samples, 18.6% (95% CI: 9.74-32.62) were positive, and C. burnetii was detected in 15% (95% CI: 8.1-26.11) of cattle milk samples. Molecular evidence of the presence of C. burnetii was seen in milk samples of dairy animals in all the studied regions. These findings demonstrated that C. burnetii infection, especially in raw milk samples, deserves more attention from the health care system and veterinary organization in Iran.

Isolation and immunogenicity of extracted outer membrane vesicles from Pseudomonas aeruginosa under antibiotics treatment conditions.

BACKGROUND AND OBJECTIVES: Different types of antibiotics have been indicated to enhance the secretion of OMVs from Pseudomonas aeruginosa. We aimed to investigate the effect of meropenem and amikacin antibiotics on inducing the secretion of OMVs and immunologic features in P. aeruginosa. MATERIALS AND METHODS: The OMVs were prepared from P. aeruginosa under hypervesiculation condition (treatment with amikacin and meropenem), and extraction was carried out by the sequential ultracentrifugation. Physicochemical features of extracted OMVs were evaluated by electron microscopy and SDS-PAGE. To quantify antibody synthesis and function after immunization with OMV, we used ELISA, serum bactericidal activity, and opsonophagocytosis. Production of cytokines from splenocytes of immunized mice was measured with ELISA. RESULTS: Specific-antibody IgG production, particularly IgG1 subclass, increased in mice primed with hypervesiculation-derived OMVs compared to normal condition-derived OMVs. Serum bactericidal activity and opsonophagocytosis of secreted antibody was enhanced in mice primed with hypervesiculation-derived OMVs. Investigation of cytokine production showed the upregulation of IL-8, IL-12, IL-17, and TNF-α and downregulation of IL-10. CONCLUSION: Based on our findings, OMVs production can be increased by treating P. aeruginosa with amikacin and meropenem antibiotics. Moreover, hypervesiculation-derived OMV scan possibly activate the humoral and cellular immune response more than normal OMVs.

A novel method for the extraction of outer membrane vesicles (OMVs) from Bordetella pertussis Tohama strain.

BACKGROUND AND OBJECTIVES: There are many pertussis outbreaks which is mainly due to the reduction in the immunity of acellular pertussis (aP) vaccines. Therefore, there is a crucial necessity to develop a new generation of pertussis vaccine. Preceding researches have shown that Bordetella pertussis outer membrane vesicles (OMVs) have appropriate specifications, making them a suitable vaccine candidate against pertussis. MATERIALS AND METHODS: The OMVs were separated by a new serial ultra centrifugation technique. Transmission electron microscopy (TEM) examination, SDS-PAGE, Western blotting and ELISA assay were used to characterize the OMVs. RESULTS: TEM studies showed the size of the extracted OMVs at 40-200 nm. The presence of pertussis toxin, filamentous hemagglutinin, and pertactin was verified using Western blot and ELISA assay. CONCLUSION: The presented technique is a simple and effective way to obtain OMVs from Bordetella pertussis. So it can be utilized as an appropriate procedure in the development of an OMV-based vaccine against pertussis.

Induction of protective immunity by recombinant peptidoglycan associated lipoprotein (rPAL) protein of Legionella pneumophila in a BALB/c mouse model.

Legionella pneumophila causes a severe form of pneumonia known as Legionnaires' disease especially in patients with impaired cellular immune response. In order to prevent the disease, immunogenicity and the level of the induction of protective immunity from the recombinant peptidoglycan-associated lipoprotein (rPAL) against Legionella pneumophila in BALB/c mice was examined. Mice immunized with (rPAL) rapidly increased an antibody response in serum and also displayed a strong activation of both innate and adaptive cell-mediated immunity as determined by antigen-specific splenocyte proliferation, an early production of pro-inflammatory cytokines in the serum and in the splenocyte cultures. Infection with a primary sublethal does of Legionella pneumophila serogroup 1, strain paris, caused resistance to a lethal challenge infection in the animals with 100% survival rate. However, mice treated with rPAL survived with 60% rate in 10 days after a lethal i.v challenge with L. pneumophila. All of the control animals receiving PBS died within 24 h. The present study indicates that recombinant protein PAL of Legionella pneumophila is strongly immunogenic and capable to elicit early innate and adaptive immune responses and lasting immunity against a lethal dose of Legionella pneumophila challenge. Antigenic characterization and immune protection of recombinant protein PAL would be of considerable value in comprehension the immune-pathogenesis of the disease and in development possible vaccine against the Legionella.

Prevalence of biofilm formation and vancomycin-resistant genes among Enterococcus faecium isolated from clinical and environmental specimens in Lorestan hospitals.

BACKGROUND AND OBJECTIVES: The antibiotic resistance among Enterococcus faecium strains has increased worldwide. Additionally, biofilm-forming isolates of E. faecium play an important role in human infections. This study was conducted to investigate the prevalence of virulence and antibiotic resistance genes between biofilm-producing and non-biofilm-producing E. faecium strains. MATERIALS AND METHODS: In this study, 228 E. faecium isolates from clinical and environmental specimens were obtained from different wards of hospitals in Lorestan province (Iran). Then, the pattern of antibiotic resistance and minimum inhibitory concentration (MIC) against ß-lactams, glycopeptides, aminoglycosides and other common antibiotics was investigated using disk diffusion and agar dilution methods. Biofilm formation was investigated using polystyrene microtiter plates. PCR assay was conducted for antibiotic resistance and biofilm related genes. Pulse field gel electrophoresis (PFGE) was used to determine the clonal spread of isolates. RESULTS: Most of isolates (78%) were resistant to penicillin, but all were susceptible to linezolid and tigecycline. The biofilm-producing isolates were more resistant to ß-lactams, glycopeptides and aminoglycosides compared to non-biofilm-producing strains. In biofilm-producing isolates, pilA, pilB, efaAfm and esp were the dominant virulence genes and vanA and pbp5 genes were the dominant resistant genes. PFGE analysis exhibited a similar pattern between the clinical and environmental isolates, suggesting the presence of a common origin of the infection by E. faecium. CONCLUSION: The results of the antibiotic resistance, biofilm assay, and PFGE analysis suggest that there is a common clone of persistent and biofilm-producing strains of E. faecium, which could rapidly disseminate in patients and the environment.

Binding of the Helicobacter pylori OipA causes apoptosis of host cells via modulation of Bax/Bcl-2 levels.

The H. pylori outer inflammatory protein A (OipA) is an outer membrane protein that contributes to gastric inflammation. OipA is believed to affect intra-cellular signalling and modulate the host signalling pathways. The aim of the current study was to clarify the role of OipA in H. pylori pathogenesis and its effect on host cell signalling pathways. To this end, the oipA gene was isolated and inserted into cloning and expression vectors. The recombinant plasmid was transferred into an expression host to produce OipA, which was subsequently purified by affinity chromatography and used for antibody production. A confluent monolayer of gastric cell lines was treated with various concentrations of OipA and investigated for attachment, toxicity, and apoptosis and alterations in signalling pathways. OipA bound to gastric cell lines confirming its role in the attachment of H. pylori to host cells. The ratio of Bax/Bcl-2 and caspase3, 8, FasL in the host cells were assessed and the results showed that the Bax/Bcl-2 ratio as well as the level of cleaved-caspase 3 was elevated in OipA-treated cells. These findings suggest that OipA can bind and induce toxic events as well as triggering apoptotic cascade in host gastric cells through intrinsic pathway.

Genotyping of clinical and environmental multidrug resistant Enterococcus faecium strains.

CONTEXT: Multidrug resistant (MDR) Enterococcus faecium is a nosocomial pathogen and clonal complex 17 (CC17) is the main genetic subpopulation of E. faecium in hospitals worldwide. AIMS: There has thus far been no report of major E. faecium clones in Iranian hospitals. SUBJECTS AND METHODS: The present study analyzed strains of MDR E. faecium obtained from patients and the Intensive Care Unit environments using pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) to determine the antibiotic resistance patterns and genetic features of the dominant. RESULTS: clones of E. faecium. PFGE and MLST analysis revealed the presence of 17and 15 different subtypes, respectively. Of these, 18 (86%) isolates belonged toCC17. Most strains in this clonal complex harbored the esp gene and exhibited resistance to vancomycin, teicoplanin, ampicillin, ciprofloxacin, gentamicin, and erythromycin. The MLST results revealed 12 new sequence types (ST) for the first time. Approximately 50% of the STs were associated with ST203. CONCLUSION: Detection of E. faecium strains belonging to CC17 on medical equipment and in clinical specimens verified the circulation of high-risk MDR clones among the patients and in hospital environments in Iran.

Gentamicin induces efaA expression and biofilm formation in Enterococcus faecalis.

Enterococci have been ranked among the leading causes of nosocomial bacteremia and urinary tract infection. This study aimed to investigate the effect of ampicillin, vancomycin, gentamicin and ceftizoxime on biofilm formation and gene expression of colonization factors on Enterococcus faecalis. Twelve clinical isolates of E. faecalis were used to investigate the effect of antibiotics on biofilm formation and gene expression of efaA, asa1, ebpA, esp and ace. Flow system assay and Microtiter plates were used for biofilm assay. Two hundred clinical isolates were used for confirming the effect of antibiotics on biofilm formation. Ampicillin, vancomycin and ceftizoxime did not have any significant effect on biofilm formation, but gentamicin induced biofilm formation in 89% of isolates. In twelve selected isolate gentamicin increased expression of esp (+50.9%) and efaA (+33.9%) genes and reduced or maintained expression of others (asa1:-47.4%, ebpA: 0, ace:-19.2%). Vancomycin increased expression of esp (+89.1%) but reduced the others (asa1: -34.9%, ebpA:-11%, ace:-30%, efaA:-60%). Ceftizoxime increased slightly ebpA (+19.7%) and reduced others (asa1:-66.2%, esp:-35%, ace:-28.1%, efaA:-38.4%). and ampicillin strongly increased expression of ace (+231%), esp (+131%) and ebpA (+83%) but reduced others (asa1:-85.5%, efaA:-47.4%). The findings of the present study showed that antibiotics may have a role in biofilm formation and sustainability of enterococci, especially in case of gentamicin. efaA gene may have an important role, especially in antibiotic induced biofilm formation by gentamicin. Experiments with efaA mutants are needed to investigate the exact effect of efaA on biofilm formation with antibiotic induced cells.

Production of specific IgY Helicobacter pylori recombinant OipA protein and assessment of its inhibitory effects towards attachment of H. pylori to AGS cell line.

PURPOSE: The common triple therapy for Helicobacter pylori is challenged by the increasing cases of antibiotic resistant infections, raising the need to explore alternative therapies. Oral administration of egg yolk immunoglobulin Y (IgY) has been previously reported as a means of passive immunization therapy for H. pylori infections. In this work, we investigated the inhibitory effect of IgY on the attachment of H. pylori to AGS cell line. MATERIALS AND METHODS: Recombinant OipA was prepared. Hens were immunized with recombinant protein three times. IgY was purified from egg yolks of immunized hens using polyethylene glycol precipitation method. The inhibitory effect of the specific immunoglobulin was evaluated in AGS cell line infected with H. pylori. RESULTS: The presence of recombinant OipA (30 kD) was confirmed via sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Immunization of hens was confirmed using enzyme-linked immunosorbent assay. The purified IgY from egg yolks were assessed using SDS-PAGE and confirmed by western blot. CONCLUSION: The results showed that IgY-OipA had inhibitory effect on attachment of H. pylori to AGS cell line and may be utilized as a therapeutic or prophylaxis material.

In silico analysis for identifying potential vaccine candidates against Staphylococcus aureus.

PURPOSE: Staphylococcus aureus is one of the most important causes of nosocomial and community-acquired infections. The increasing incidence of multiple antibiotic-resistant S. aureus strains and the emergence of vancomycin resistant S. aureus strains have placed renewed interest on alternative means of prevention and control of infection. S. aureus produces a variety of virulence factors, so a multi-subunit vaccine will be more successful for preventing S. aureus infections than a mono-subunit vaccine. MATERIALS AND METHODS: We selected three important virulence factors of S. aureus, clumping factor A (ClfA), iron-regulated surface determinant (IsdB), and gamma hemolysin (Hlg) that are potential candidates for vaccine development. We designed synthetic genes encoding the clfA, isdB, and hlg and used bioinformatics tools to predict structure of the synthetic construct and its stabilities. VaxiJen analysis of the protein showed a high antigenicity. Linear and conformational B-cell epitopes were identified. RESULTS: The proteins encoded by these genes were useful as vaccine candidates against S. aureus infections. CONCLUSION: In silico tools are highly suited to study, design, and evaluate vaccine strategies.

Concomitant Colonization of Helicobacter pylori in Dental Plaque and Gastric Biopsy.

Frequently reported H. pylori antimicrobial therapy failures suggest that there might be a different niche where the bacteria can stay safe. Current study aims to examine potential role of oral colonization of H. pylori to feed reinfection after primary therapy. However, patients who were admitted to the gastroscopy section were chosen and gastric biopsy and dental plaque specimens were collected. Molecular and biochemical tests were applied to confirm H. pylori identity in different colonization niches. Results showed that 88.8% of dyspeptic patients had epigastric pains with nocturnal awakening when they were hungry (P = 0.023). All patients who received therapy already were again H. pylori positive while they are still carrying H. pylori in dental plaque (P = 0.001). Moreover, H. pylori infection was sought in 100% of gastric biopsy's dyspeptic patients who had ulcerated esophagitis and erosive duodenitis and who were H. pylori positive, and 75% of dyspeptic patients with duodenum deformity had this bacterium in gastric biopsies (P = 0.004). Present study showed that only successful eradication of gastric H. pylori cannot guarantee prevention of reinfection. Conclusively, a new strategy which indicates concomitant eradication in oral and gastric colonization can result in clearance of H. pylori infection.

Clinical relevance of cagL gene and virulence genotypes with disease outcomes in a Helicobacter pylori infected population from Iran.

Helicobacter pylori infection is common in Iran as in other developing countries. Certain genotypes of H. pylori have been associated with increased occurrence of chronic gastritis, peptic ulcers, and gastric adenocarcinoma. The aim of this study was to investigate the clinical relevance of cagL gene and other virulence genotypes of H. pylori isolates with clinical outcomes in Iranian patients. Totally, 126 symptomatic patients who underwent gastroduodenal endoscopy were enrolled in the study. Sixty-one H. pylori strains were isolated from the patients studied. The presence of the cagL, cagA, vacA, iceA, babA2 and sabA genes in the corresponding H. pylori isolates were determined by polymerase chain reaction and the results were compared with clinical outcomes and histopathology. The cagL, cagA, vacA s1, vacA s2, vacA m1, vacA m2, iceA1, iceA2, babA 2 , and sabA genotypes were detected in 96.7, 85.2, 75.4, 24.6, 29.5, 70.5, 42.6, 23, 96.7, and 83.6% of the isolates, respectively. The three genotypic combinations, cagL/cagA/vacAs1m1/iceA1/babA2/sabA, cagL/cagA/vacAs1m2/iceA1/babA2/sabA, and cagL/cagA/vacAs1m2/iceA2/babA2/sabA were determined as the most prevalent combined genotypes. There was a significant correlation between the presence of cagL gene and cagA positivity (P = 0.02). No significant correlation was found between the various genotypes and clinical outcomes (P > 0.05). The present study showed a very high prevalence of cagL genotype among the H. pylori isolates from Iranian patients. Our results demonstrated that neither single genotype nor combination genotypes of virulence-associated genes was significantly helpful markers for predicting the severity of gastroduodenal disease associated with H. pylori infection in Iranian patients.

High frequency distribution of heterogeneous vancomycin resistant Enterococcous faecium (VREfm) in Iranian hospitals.

BACKGROUND: Enterococcus faecium is a multi-resistant nosocomial pathogen causing infection in debilitated patients. Vancomycin-resistant enterococcus faecium (VREfm) are a major concern and increased dramatically worldwide especially in hospitals environment. The current study focused on determining the high prevalence and distribution patterns of antibiotic resistance and also its genetic linkages among various VREfm strains isolated from indoor hospitalized patients in four major Iranian teaching hospitals of Tehran. METHODS: The clinical samples were obtained from hospitalized patients during September 2010 to June 2011 from different teaching hospitals of Tehran. Antibiotics Resistance patterns, minimum inhibition concentration (MIC) value for vancomycin, ampicillin, gentamicin and presence of genetic linkage among the isolates were determined by pulsed-field gel electrophoresis (PFGE). RESULTS: Overall, total of 92 (41.4%) isolates were identified as E. faecium, 45 (49%) were resistant to vancomycin with an MIC50 of ≥ 128 mg/L. The results showed that simultaneous resistance to teicoplanin, ampicillin, gentamicin, ciprofloxacine, tetracycline and erythromycin were observed the most frequent pattern. All the vancomycin resistant E. faecium isolates carried the vanA gene. intensive care units (ICUs) and Kidney transplantation, are most probably the wards with highest risk of infection by VRE. 17 pulsotypes were also detected by PFGE, most of the related pulsotypes belongs to the same hospitals. CONCLUSIONS: This study shows the high alarming prevalence of Enterococcus faecium infection and similar clones of VREfm strains in Iranian hospitals with threatening resistance phenotypes. VIRTUAL SLIDES: The virtual slides for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1270863903102282.

Adhesion and virulence factor properties of Enterococci isolated from clinical samples in Iran.

INTRODUCTION: Enterococci rank among leading causes of nosocomial bacteremia, urinary tract infections and community acquired endocarditis. The aim of the present study was to investigate the presence of virulence factors in Enterococci strains isolated from clinical samples in Iranian Educational hospitals. METHODOLOGY: Presence of aggregation substance (asa), extracellular surface protein (esp), Enterococcus faecalis antigen A (efaA), adhesin of collagen from E. faecalis (ace), endocarditis and biofilm-associated pilli (ebp) as colonization factors and cytolysin (cyl), gelatinase (gel) and hyaloronidase (hyl) as secretary factors were investigated in isolates. A total of 201 clinical isolates of Enterococci were collected in 2009-2010 from eight educational hospitals. After deoxyribonucleic acid extraction, they were examined for presence of virulence factors by polymerase chain reaction. RESULTS: E. faecalis and Enterococcus faecium were isolated from 56.9% to 43.1%, respectively. Resistance to vancomycin and gentamicin were 33.8% and 83.9% in E. faecium isolates and 16.3% and 88.1% in E. faecalis isolates respectively. Colonization factors were found to be more prevalent in E. faecalis isolates and almost all isolates of E. faecalis had ace, ebp and efaA genes. Esp gene had a higher rate of distribution in Enterococci isolates (75.1%) in this study compared with previous studies. One of E. faecalis isolates contained hyl gene, but 38.8% of E. faecium isolates had it. Mutual exclusive were present between hyl and efaA in all E. faecium isolates and 69.7% of E. faecium hyl - positive isolates were esp positive. CONCLUSION: According to these results, virulence genes were more prevalent in E. faecalis isolates and E. faecalis had more potential pathogenesis for initiating an infection; however because of E. faeciums higher antibiotic resistance, we have been facing higher E. faecium infections in hospitalized patients.

In silico experiment with an-antigen-toll like receptor-5 agonist fusion construct for immunogenic application to Helicobacter pylori.

BACKGROUNDS: Helicobacter pylori colonize the gastric mucosa of half of the world's population. Although it is classified as a definitive type I carcinogen by World Health Organization, there is no effective vaccine against this bacterium. H. pylori evade the host immune response by avoiding toll-like detection, such as detection via toll-like receptor-5 (TLR-5). Thus, a chimeric construct consisting of selected epitopes from virulence factors that is incorporated into a TLR-5 ligand (Pseudomonas flagellin) could result in more potent innate and adaptive immune responses. MATERIALS AND METHODS: Based on the histocompatibility antigens of BALB/c mice, in silico techniques were used to select several fragments from H. pylori virulence factors with a high density of B- and T-cell epitopes. RESULTS: These segments consist of cytotoxin-associated geneA (residue 162-283), neutrophil activating protein (residue 30-135) and outer inflammatory protein A (residue 155-268). The secondary and tertiary structure of the chimeric constructs and other bioinformatics analyses such as stability, solubility, and antigenicity were performed. The chimeric construct containing antigenic segments of H. pylori proteins was fused with the D3 domain of Pseudomonas flagellin. This recombinant chimeric gene was optimized for expression in Escherichia coli. The in silico results showed that the conserved C- and N-terminal domains of flagellin and the antigenicity of selected fragments were retained. DISCUSSION: In silico analysis showed that Pseudomonas flagellin is a suitable platform for incorporation of an antigenic construct from H. pylori. This strategy may be an effective tool for the control of H. pylori and other persistent infections.

Bioactivity and immunological evaluation of LPS from different serotypes of Helicobacter pylori.

BACKGROUND AND OBJECTIVES: Helicobacter pylori is the causative agent of peptic ulcer disease and a co-factor in development of gastric malignancies. LPS are among toxic substances produced by H. pylori exhibiting low endotoxic activity compared to typical bacterial LPS. The aim of this study was to investigate bioactivity of LPS produced by different serotypes of Helicobacter pylori compared to Escherichia coli and Brucella abortus LPS. MATERIALS AND METHODS: Bacterial LPS was extracted by the hot phenol-water method. Biological activities of LPS were determined via the limulus lysate assay, pyrogenic assay, and blood pressure and PBMC induction test in rabbits. RESULTS: Biological activity of O2 serotype LPS of H. pylori was less than the biological activity of other H. pylori serotypes. CONCLUSION: Our data supported the hypothesis that the unique bacterial LPS of the O2 serotype must be included in the formulation of a multivalent H. pylori vaccine.

Cloning, Expression, Purification and Toxicity Evaluation of Helicobacter pylori Outer Inflammatory Protein A.

The Helicobacter pylori outer membrane proteins play an important role in pathogenesis; the outer inflammatory protein A (OipA) is one of these proteins which play the main role in the development of inflammation. In this study, purification of recombinant H. pylori OipA was performed by Ni-NTA affinity chromatography. Gastric carcinoma epithelial cells (AGS cell) were treated by different concentrations of recombinant OipA for various lengths of time and cell viability was evaluated by the viability assay. Statistical analysis showed that OipA had toxic effects on AGS cells in a concentration of 500 ng/ml after 24 and 48 h, and this toxic dose was 256 ng/ml after 72 h. OipA had direct toxic effects on gastric epithelial cells and the toxicity was observed to depend on time and dose of H. pylori exposure. Attachment of H. pylori to gastric epithelial cells is a key part in the pathogenesis and enables H. pylori to damage the epithelial cells with OipA.

Colonization of hospital water systems by Legionella pneumophila, Pseudomonas aeroginosa, and Acinetobacter in ICU wards of Tehran hospitals.

BACKGROUND: Nosocomial infection caused by non-Enterobacteriaceae gram negative bacteria (GNB-NE) is increasing in intensive care units (ICU). AIM: The objective of this study was to determine whether potable water in ICU wards at Tehran hospitals is contaminated with L. pneomophila, P. aeroginosa and Acinetobacter spp. MATERIALS AND METHODS: A total of 52 water samples from shower bath and taps water in seven hospitals of Tehran were collected. The water sample concentrated by filtering through millipore cellulose filters and cultured on BCYE agar and tryptic soya agar media. The presence of Legionella pneumophila was confirmed by real time PCR assay using primers-probe designed for the mip gene. RESULTS: Legionella pneumophila, Pseudomonas aeroginosa and Acinetobacter were isolated from 5 (9.6%), 6 (11.4%) and 1 (1.8%) of the hospital water systems, respectively. This study demonstrated the presence of Legionella, Pseudomonas and Acinetobacter in water system in ICU wards of different hospitals in Tehran. CONCLUSIONS: Hot water from shower heads could be a potential source of infection for Legionella pneumophila. Water was also proved to contain Pseudomonas aeruginonsa, the main GNB-NE causing nosocomila pneumonia at Tehran hospitals. Care should be taken concerning cleanliness and decontamination of water supplies at ICUs for pathogenic organisms.