Antimicrobial, anti-adhesive, and anti-invasive effects of condition media derived from adipose mesenchymal stem cells against Shigella flexneri.

The objective of the current study was to examine the antimicrobial, anti-adhesion, and anti-invasion properties of various concentrations of condition media obtained from adipose mesenchymal stem cells (AD-MSCs CM) against Shigella flexneri (S. flexneri). AD-MSCs characterization and antimicrobial assay were performed using flow cytometry and microdilution by colony counting, respectively. For evaluating adhesion and invasion, Caco-2 cells were infected by S. flexneri at three different multiplicities of infection (MOIs of 1, 10, and 50) and then treated with DMEM medium and AD-MSCs CM. The inhibitory effect of AD-MSCs CM was assessed after 24 and 48 h of treatment by CFU (colony-forming unit) counting. A total of 84, 65, and 56% reduction in the adhesion rate of S. flexneri to Caco-2 cells treated with AD-MSCs CM were observed at MOIs of 1, 10, and 50, respectively. While S. flexneri at MOI:1 had no invasive effect on Caco-2 cells, convincing invasion was detected at MOIs of 10 and 50, showing a significant decrease following treatment with AD-MSCs CM. The current study results open new insights into AD-MSCs CM as a new non-antibiotic therapeutic candidate for S. flexneri infections.

Design of dot-blot hybridization assay for simultaneous detection of Campylobacter jejuni and Campylobacter coli: a preliminary study.

Objectives: Campylobacters are a major cause of gastroenteritis worldwide. These are fastidious in culture and false negative results are seen in many clinical laboratories. Among molecular methods, the dot-blot technique is widely used for a variety of purposes, especially diagnostics. So, the authors aimed to detect C. jejuni and C. coli simultaneously using a dot-blot assay. Methods: After evaluating the bioinformatics studies, a cadF-conserved fragment was selected for the design of primers and probe. DNAs from standard strains and a recombinant plasmid, prepared in this study, were used to assess the technique. The specificity of the method was also surveyed using DNAs from other enteric bacteria. The limit of detection was evaluated by recombinant plasmid and different concentrations of the designed probe. Results: A 95-bp fragment of cadF was selected, and in silico analysis studies showed that it is conserved between both species. Also, the non-specific annealing of the primers and probe with other bacteria was not seen theoretically. The technique with recombinant plasmid as well as DNAs of standard strains created black spots on the membrane, confirming that the probe was correctly synthesized. No non-specific reactions with other bacterial species were observed (specificity=100%). The limit of detection of the test was determined to be 50 µg/ml. Conclusions: This is the first study to simultaneously detect two important pathogens in the Campylobacter genus and was able to detect C. jejuni and C. coli with acceptable sensitivity and specificity.

Genetic approach toward linkage of Iran 2012-2016 cholera outbreaks with 7th pandemic Vibrio cholerae.

Vibrio cholerae, as a natural inhabitant of the marine environment is among the world-leading causes of diarrheal diseases. The present study aimed to investigate the genetic relatedness of Iran 2012-2016 V. cholerae outbreaks with 7th pandemic cholera and to further characterize the non-ST69/non-ST75 sequence types strains by whole-genome sequencing (WGS).Twenty V. cholerae isolates related to 2012, 2013, 2015 and 2016 cholera outbreaks were studied by two genotyping methods - Pulsed-field Gel Electrophoresis (PFGE) and Multi-locus Sequence Typing (MLST)-and by antimicrobial susceptibility testing. Seven sequence types (STs) and sixteen pulsotypes were detected. Sequence type 69 was the most abundant ST confirming that most (65%, 13/20) of the studied isolates collected in Iran between 2012 and 2016 belonged to the 7th pandemic clone. All these ST69 isolates (except two) exhibited similar pulsotypes. ST75 was the second most abundant ST. It was identified in 2015 and 2016. ST438, ST178, ST579 and STs of 983 and 984 (as newfound STs) each were only detected in one isolate. All strains collected in 2016 appeared as distinct STs and pulsotypes indicative of probable different originations. All ST69 strains were resistant to nalidixic acid. Moreover, resistance to nalidixic acid, trimethoprim-sulfamethoxazole and tetracycline was only observed in strains of ST69. These properties propose the ST69 as a unique genotype derived from a separate lineage with distinct resistance properties. The circulation of V. cholerae ST69 and its traits in recent years in Iran proposes the 7th pandemic strains as the ongoing causes of cholera outbreaks in this country, although the role of ST75 as the probable upcoming dominant ST should not be ignored.Genomic analysis of non-ST69/non-ST75 strains in this study showed ST579 is the most similar ST type to 7th pandemic sequence types, due to the presence of wild type-El Tor sequences of tcpA and VC-1319, VC-1320, VC-1577, VC-1578 genes (responsible for polymyxin resistance in El Tor biotype), the traits of rstC of RS1 phage in one strain of this ST type and the presence of VPI-1 and VSP-I islands in ST579 and ST178 strains. In silico analysis showed no significant presence of resistance genes/cassettes/plasmids within non-ST69/non-ST75 strains genomes. Overall, these data indicate the higher susceptibility of V. cholerae non-ST69/non-ST75 strains in comparison with more ubiquitous and more circulating ST69 and ST75 strains.In conclusion, the occurrence of small outbreaks and sporadic cholera cases due to V. cholerae ST69 in recent years in Iran shows the 7th pandemic strains as the persistent causes of cholera outbreaks in this country, although the role of ST75 as the second most contributed ST should not be ignored. The occurrence of non-ST69/non-ST75 sequence types with some virulence factors characteristics in border provinces in recent years is noteworthy, and further studies together with surveillance efforts are expected to determine their likely route of transport.

Bacterial biomarkers: new aspects of colorectal tumors diagnosis: reality or fantasy.

As one of the most common cancers, colorectal cancer (CRC) is considered as a significant leading cause of cancer-related deaths worldwide. Gut microbiota is part of a complex microbial-based ecosystem in the human body so that changes in the microbiota could lead to a variety of diseases. A growing number of studies have shown that bacteria are both individually and collectively involved in the progression of CRC. The present review study provided a summary of some of the available data on the advantages and limitations of current CRC screening methods as well as gut biomarkers including genetic, epigenetic, and protein markers. Moreover, a summary of the applications and limitations in the detection of gut microbiota markers as well as their role in early diagnosis and timely treatment response in CRC patients was provided.

Characteristics of multiresistant Pseudomonas aeruginosa isolates from burn patients in Iran.

Infections caused by multidrug resistant (MDR) Pseudomonas aeruginosa isolates in burn patients restrict therapeutic strategies. The current study aimed to analyze antibiotic resistance genes and multilocus sequence typing (MLST) of P. aeruginosa strains isolated from burn patients in Shahid Motahari hospital in Tehran, Iran.Altogether 63 P. aeruginosa isolates were characterized in this study. Antibiotic susceptibility testing was performed by disc diffusion method. PCR was performed to determine the frequency of resistance genes. The expression rates of mexB, mexY genes were evaluated by Real-Time PCR. Genotyping of isolates was performed by MLST analysis. All isolates were MDR in this study. The highest resistance was detected against gentamicin, tobramycin, and cefoxitin (100%), while all isolates were susceptible to colistin. Altogether 14 resistance profiles were determined, and profile 1 included more than 50% of the isolates with the highest resistance. In this study blaampC, blaVIM-2, blaOXA-10, and aac(6')-Ib resistance genes were detected in all isolates. The expression levels of mexB and mexY genes were upregulated in 66.6 and 88.8% of MDR isolates, respectively. Overexpression of both genes was detected in 55.5% of the isolates.MLST analysis revealed five sequence types (STs), including ST235, ST664, ST532, ST2637, and ST230, which showed a significant relationship with antibiotic resistance profiles. The present study indicates an increase in antibiotic resistance against different antibiotic families among P. aeruginosa isolates. We describe the circulation of globally distributed STs among hospitalized patients, and we report ST235 as the most common MDR clone in our study.

Oxygen-Rich Graphene/ZnO2-Ag nanoframeworks with pH-Switchable Catalase/Peroxidase activity as O2 Nanobubble-Self generator for bacterial inactivation.

The oxygen-rich organic/inorganic (reduced graphene oxide (rGO)/ZnO2-Ag) nanoframeworks as suppliers of O2 nanobubbles (NBs) with dual pH-and-temperature-sensitive behavior were developed to suppress bacterial growth. It was demonstrated that not only the rate but also the final product of oxygen-rich ZnO2 decomposition (to an intermediate product of H2O2) rate was dramatically controlled by pH adjustment. Furthermore, in the presence of Ag nanoparticles, ̇OH radical generation switched to O2 NBs evolution by shifting the pH from acidic to basic/neutral conditions, demonstrating an adjustable nanozyme function-ability between catalase and peroxidase-like activity, respectively. Antibacterial properties of the in-situ generated O2 NBs substantially enhanced against bacterial models including methicillin-resistant Staphylococcus aureus in the presence of rGO. In fact, deflecting the electrons from their main respiratory chain to an oxygen-rich bypath through rGO significantly stimulated reactive oxygen species (ROS) generation, combating bacteria more efficiently. Moreover, NIR laser irradiation-induced temperature rise (due to the inherent photothermal properties of rGO) facilitated ZnO2 decomposition and accelerated growth and collapse of NBs. The simultaneous microscale thermal and mechanical destructions induced stronger antibacterial behavior. These results hold great promises for designing simple organic/inorganic nanoframeworks as solid sources of NBs with tunable enzyme-like ability in response to environmental conditions suitable for forthcoming graphene-based bio-applications.

Lytic potential of a filamentous bacteriophage isolated from sewage water in Tehran on clinical carbapenem-resistant strains.

Background and Objectives: Acinetobacter baumannii (A. baumannii), an opportunistic pathogen, has been isolated from sewage, soil, and hospital wards. The prevalence of multidrug-resistance A. baumannii has seriously caused a health crisis in hospital settings. Bacteriophages have been used as an alternative therapy for control carbapenem-resistant bacteria-caused infections. We aimed to assay lytic effect of a filamentous phage on clinical bacterial strains. Materials and Methods: 500-ml water samples was collected from sewage in Tehran. Sewage samples were precipitated at 6000 rpm for 10 minutes and filtered using 0.45-µm syringe filters. Bacteriophage was isolated using double-layer agar assay and evaluated its stability at various pH and temperature ranges. In addition, the stability of the phage was assayed at chloroform 0.1%. Results: Transmission electron microscopy imagine showed phage is filamentous called vB-AbaI-TMU2. The phage affected on its own host so that could not effect on any Pseudomonas aeruginosa and Klebsiella pneumoniae strains. vB-AbaITMU2 phage was stable at pH 5 and 7 and also temperatures 25 and 37°C. vB-AbaI-TMU2 was stable at chloroform 0.1%. vB-AbaI-TMU2 phage infected carbapenem-resistant A. baumannii strains while other bacterial strains were resistant to. Conclusion: The present study indicated the isolated phage had a narrow host range and is susceptible to various pH values and temperatures.

Mesoporous silica nano-adjuvant triggers pro-inflammatory responses in Caco-2/peripheral blood mononuclear cell (PBMC) co-cultures.

The aim of this study was to evaluate the cytotoxicity and immune-stimulatory effect of Mesoporous silica nanoparticle (MSN) Nano-adjuvant on pro-inflammatory cytokines and pattern recognition receptors (PRR) genes expression in Caco-2/PBMC co-culture model. MSNs were synthesized and characterized by scanning electron microscope (SEM), Brunauer Emmett Teller (BET) and Barrett Joyner Halenda (BJH) techniques. The BET specific surface area of MSNs was around 947 m2/g and the total pore volume and average pore diameter were 1.5 cm3/g and 8.01 nm, respectively. At the concentration of 10 µg/mL, MSN showed a low and time-dependent cytotoxicity on Caco-2 cells, while no cytotoxic effect was observed for 0.1 and 1 µg/mL concentrations after 24, 48 and 72 h. The expression of pro-inflammatory cytokines genes (IL-1, IL-8 and TNF-α) in co-cultures treated with different concentrations of MSN showed a dose-dependent significant increase up to 17.44, 2.722 and 4.34 folds, respectively, while the expression augmentation of IL-1 gene was significantly higher than the others. This indicates slight stimulation of intestinal inflammation. Different concentrations of MSN significantly increased TLR4 and NOD2 expression to 4.14 and 2.14 folds, respectively. NOD1 was not affected significantly. It can be concluded that MSN might increase protective immune responses against antigens as a vaccine adjuvant candidate. It seems that stimulation of TNF-α, IL-1, and IL-8 expression in enterocytes probably transpires through the agonistic activity of MSN for TLRs including TLR4, while NOD2-associated signaling pathways are also involved. This study provides an overall picture of MSN as a novel and potent oral adjuvant for mucosal immunity.

Effects of chitosan nanoparticles loaded with mesenchymal stem cell conditioned media on gene expression in Vibrio cholerae and Caco-2 cells.

Vibrio (V.) cholerae forms a pellicle for self-defense in the pathological conditions in the intestine, which protects it against antibiotics and adverse conditions. Targeting biofilm genes and Toll-like receptors (TLRs) is one of the new strategies to combat multidrug-resistant bacteria. The objective of this study was to evaluate the effect of mesenchymal stem cell conditioned media (MSC CM; 1000 µg), chitosan nanoparticles incorporated with mesenchymal stem cell conditioned media (MSC CM-CS NPs; 1000 µg + 0.05%), and chitosan nanoparticles (CS NPs; 0.05%) on the expression of bap1 and rbmC biofilm genes in V. cholerae and TLR2 and TLR4 genes in Caco-2 cells. The bacteria were inoculated in the presence or absence of MSC CM, MSC CM-CS NPs, and CS NPs for 24 h at 37 °C to evaluate the expression of biofilm genes. The Caco-2 cells were also exposed to V. cholerae for 1 h and then MSC CM, MSC CM-CS NPs, and CS NPs for 18 h at 37 °C. After these times, RNA was extracted from Caco-2 cells and bacteria exposed to the compounds, and the expression of target genes was evaluated using real-time PCR. Caco-2 cell viability was also assessed by MTT assay. After adding MSC CM, MSC CM-CS NPs, and CS NPs to V. cholerae medium, the percentage reduction in gene expression of bap1 was 96, 91, and 39%, and rbmC was 93, 92, and 32%, respectively. After adding MSC CM, MSC CM-CS NPs, and CS NPs to the Caco-2 cell medium, the percentage reduction in the gene expression of TLR4 was 89, 90, and 82%, and TLR2 was 41, 43, and 32%, respectively. MTT showed that Caco-2 cell viability was high and the compounds had little toxicity on these cells. Finally, it suggests that MSC CM-CS NPs designed may be a therapeutic agent to combat inflammation and biofilm formation in multidrug-resistant V. cholerae. However, further studies in vivo are also recommended.

Phenotypic and genotypic changes of Staphylococcus aureus in the presence of the inappropriate concentration of chlorhexidine gluconate.

BACKGROUND: Chlorhexidine gluconate (CHG) is a disinfectant agent with different applications in health care. Improper use of CHG causes antimicrobial resistance in bacteria as a public health threat. Since Staphylococcus aureus is a common bacteria, it is expected usually exposed to CHG in the hospital and community. The present study aimed to correlate the phenotypic and genotypic changes in a S. aureus strain upon serial adaptation with supra-inhibitory CHG concentration for 50 days. RESULTS: After in vitro serial culture of 5 × 105 CFU/ml of a clinical vancomycin-susceptible S. aureus strain (VAN-S) into brain heart infusion (BHI) broth containing CHG 1/4, 1/2, 1, and 2 × minimal inhibitory concentration (MIC) values of VAN-S in 37 °C during 50 days, we isolated a S. aureus strain (CHGVan-I) with a ≥ twofold decrease in susceptibility to CHG and vancomycin. CHG-induced CHGVan-I strain was considered as a vancomycin-intermediate S. aureus (VISA) strain with a VAN MIC of 4 µg/ml using the broth macro dilution method. However, reduced resistance was observed to tetracycline family antibiotics (doxycycline and tetracycline) using a modified Kirby-Bauer disk diffusion test. Moreover, a remarkable reduction was detected in growth rate, hemolysis activity (the lysis of human red blood cells by alpha-hemolysin), and colony pigmentation (on BHI agar plates). Biofilm formation (using the Microtiter plate method and crystal violet staining) was significantly increased upon CHG treatment. Adaptive changes in the expression of a set of common genes related to the development of VISA phenotype (graTSR, vraTSR, walKR, agr RNAIII, sceD, pbpB, and fmtA) were analyzed by Reverse Transcription quantitative PCR (RT-qPCR) experiment. Significant changes in vraTSR, agr RNAIII, sceD, and pbpB expression were observed. However, gene sequencing of the two-component system vraTSR using the Sanger sequencing method did not detect any non-synonymous substitution in CHGVan-I compared to wild-type. The clonality of VAN-S and CHGVan-I strains was verified using the pulsed-field gel electrophoresis (PFGE) method. CONCLUSIONS: The importance of the present study should be stated in new detected mechanisms underlying VISA development. We found a link between the improper CHX use and the development of phenotypic and genotypic features, typical of VISA clinical isolates, in a CHG-induced strain. Since disruption of the cell wall biosynthesis occurs in VISA isolates, our CHG-induced VISA strain proved new insights into the role of CHG in the stimulation of the S. aureus cell wall.

Investigation of the Antibacterial and Antibiofilm Activity of Selenium Nanoparticles against Vibrio cholerae as a Potent Therapeutics.

Vibrio cholerae is a major cause of severe diarrhea, which is ecologically flexible, and remains as a major cause of death, especially in developing countries. Consecutive emergence of antibiotic-resistant strains is considered to be as one of the major concerns of the World Health Organization (WHO). Nanoparticles as a new nonantibiotic therapeutic strategy have been widely used in recent years to treat bacterial infections. The present study aimed to investigate the antibacterial and antibiofilm effect of selenium nanoparticles (SeNPs) in vitro against V. cholerae O1 ATCC 14035 strain. SeNPs were prepared and characterized using ultraviolet-visible (UV-Vis) spectroscopy, DLS (dynamic light scattering), zeta potential measurement, and Fourier transform infrared (FTIR) analysis. The concentration of SeNPs was calculated by ICP (inductively coupled plasma) method. Also, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was employed to assess the cytotoxic effect of SeNPs on Caco-2 cells. Antibacterial and antibiofilm activity of SeNPs was determined by broth microdilution and crystal violet assays, respectively. The average particle size of SeNPs was 71.1 nm with zeta potential -32.2 mV. The SEM images supported the uniform spherical morphology of the prepared nanoparticles. The antibiofilm effect of SeNPs was evident at concentrations of 50-200 µg/mL. This study results provided evidence that SeNPs are safe as an antibacterial and antibiofilm agent against V. cholerae O1 ATCC 14035 strain.

Formulation of selenium nanoparticles encapsulated by alginate-chitosan for controlled delivery of Vibrio Cholerae LPS: A novel delivery system candidate for nanovaccine.

The lipopolysaccharide (LPS) of Vibrio cholerae plays a significant role in stimulating primary protection and immune responses. LPS delivery has been limited by the stimulation of inflammatory cytokines. This work aimed to report the synthesis and performance of this formulation in modulating immune responses and protecting LPS against acidic gastric medium. Alg-Cs-LPS-SeNPs composite was fabricated by an ionic cross-linking/in situ reduction method. Cytokines TNF-α, IL-6, IL-10, and TGF-ß were assessed after cells were incubated with different compounds of the system. The main outcomes revealed that encapsulation of LPS-loaded SeNPs in the alginate-chitosan complex was associated with a high entrapment efficiency and could effectively protect LPS against acidic GIT medium. Kinetic profiling revealed that LPS was more slowly released from LPS-loaded Alg-Cs-LPS-SeNPs at pH 1.2, 7.4, and 6.8. These results indicated that Alg-Cs-LPS-SeNPs composite was able to significantly increase anti-inflammatory cytokines and reduce the release of pro-inflammatory cytokines. Thus, these findings show that this system for LPS delivery could be easily biosynthesized and encapsulated for use in the pharmaceutical industry. This study provides proof of the potential for future use of oral LPS vaccines, concomitantly inducing immunomodulatory effects.

Lifetime physical activity is associated with gut bacteria and brain health in people with multiple sclerosis: Focus on physical activity intensity.

BACKGROUND: Physical activity can improve brain health in people with multiple sclerosis (PwMS). One of the underlying mechanisms can be the modulation of gut bacteria. The association of different intensity of physical activity in a lifetime; and brain volume, lesion volume, and number, and gut bacteria counts were investigated in the current study. METHODS: Forty-five PwMS were recruited, and magnetic resonance imagining was used to evaluate brain volume, lesion volume, and number. Also, stool samples were taken for evaluation faecalibacterium prausnitzii, akkermansia muciniphila, prevotella, and bacteroides count. Moreover, lifetime physical activity was assessed using the adapted version of the historical activity questionnaire. RESULTS: Data revealed a significant association of physical activity with brain volume (r = 0.41), lesion volume (r=-0.35), lesion number (r=-0.37), akkermansia muciniphila (r=-0.34), prevotella (r = 0.52) and bacteroides (r=-0.32) count (p<0.05). Moderate-intensity of physical activity was associated with brain volume (r = 0.33), lesion volume (r=-0.38), prevotella (r = 0.35) and bacteroides (r=-0.40) count (p<0.05). Moreover, vigorous-intensity of physical activity was associated with brain volume (r = 0.38), lesion number (r=-0.39), akkermansia muciniphila (r=-0.30) and prevotella (r = 0.56) count (p<0.05). CONCLUSION: Our results suggest that lifetime physical activity is associated with brain health and gut bacteria count in PwMS. Additionally, the heterogeneity of the association of the physical activity intensities with the studied variables indicates the importance of using different intensities of physical activity to greater benefit from physical activity.

Tropheryma whipplei in the stool samples of children with acute diarrhea: a study from Tehran, Iran.

BACKGROUND: Recently, Tropheryma whipplei has been suggested as one of the causative agents of diarrhea among children worldwide. Limited data is available on the prevalence of T. whipplei among children with diarrhea in most countries such as Iran. This study was conducted to evaluate the prevalence of T. whipplei in children with acute diarrhea in Iran. METHODS: In this study, the stool samples were collected from 130 children under 10 years old with acute diarrhea from children's hospitals in Tehran city. Genomic DNA was extracted from stool samples and was tested for the presence of DNA of T. whipplei using the SYBR Green Real-time PCR method. Positive T. whipplei samples were finally confirmed by PCR Product sequencing. RESULTS: The mean age of participants was 32.5 months, and 54.6% of children were female. Using the SYBR Green Real-time PCR, 9.23% (12/130) of samples were positive for T. whipplei, which were confirmed by sequencing. 66.67% of positive cases were males. The duration of diarrhea in infected children with T. whipplei (83.3%) was significantly longer (OR: 5.93, 95% CI 1.24-28.22) compared to children with negative results (45.8%). Other demographic factors and clinical signs had not a statistically significant relationship with T. whipplei infection. CONCLUSIONS: In this study, T. whipplei was detected in stool samples of children with acute diarrhea. The results indicated that T. whipplei could be associated with childhood diarrhea in Iran. The health care system and physicians should be aware of the presence of T. whipplei infection in Iran, especially in childhood diarrhea.

Immunomodulatory effects of mesenchymal stem cell-conditioned media on lipopolysaccharide of Vibrio cholerae as a vaccine candidate.

BACKGROUND: Vibrio cholerae is the causative agent of cholera, which is commonly associated with high morbidity and mortality, and presents a major challenge to healthcare systems throughout the world. Lipopolysaccharide (LPS) is required for full protection against V. cholerae but can induce inflammation and septic shock. Mesenchymal stem cells (MSCs) are currently used to treat infectious and inflammatory diseases. Therefore, this study aimed to evaluate the immune-modulating effects of the LPS-MSC-conditioned medium (CM) on V. cholerae LPS immunization in a murine model. METHODS: After preconditioning MSCs with LPS, mice were immunized intraperitoneally on days 0 and 14 with the following combinations: LPS + LPS-MSC-CM; detoxified LPS (DLPS) + MSC-CM; LPS + MSC sup; LPS; LPS-MSC-CM; MSC supernatant (MSC sup); and PBS. The mouse serum and saliva samples were collected to evaluate antibody (serum IgG and saliva IgA) and cytokine responses (TNF-α, IL-10, IL-6, TGF-ß, IL-4, IL-5, and B-cell activating factor (BAFF)). RESULTS: The LPS + LPS-MSC-CM significantly increased total IgG and IgA compared to other combinations (P < 0.001). TNF-α levels, in contrast to IL-10 and TGF-ß, were reduced significantly in mice receiving the LPS + LPS-MSC-CM compared to mice receiving only LPS. IL-4, IL-5, and BAFF levels significantly increased in mice receiving increased doses of LPS + LPS-MSC-CM compared to those who received only LPS. The highest vibriocidal antibody titer (1:64) was observed in LPS + LPS-MSC-CM-immunized mice and resulted in a significant improvement in survival in infant mice infected by V. cholerae O1. CONCLUSIONS: The LPS-MSC-CM modulates the immune response to V. cholerae LPS by regulating inflammatory and anti-inflammatory responses and inducing vibriocidal antibodies, which protect neonate mice against V. cholerae infection.

Transcriptome analysis of biofilm formation under aerobic and microaerobic conditions in clinical isolates of Campylobacter spp.

It has been well documented that Campylobacter is the leading cause of foodborne infections and bacterial enteritis in high-income countries. The gastrointestinal tract of most warm-blooded animals, such as mammals and poultry, is prone to this pathogen. Infections caused by this bacterium in humans have usually been associated with the consumption of contaminated poultry meat. The important point about Campylobacter is that this bacterium has adapted to harsh environmental conditions along the food chain (poultry digestive tract to the consumer's plate) and developed an adapted mechanism to those conditions. This study aimed to compare the ability of Campylobacter jejuni and Campylobacter coli strains to form biofilms under aerobic and microaerobic conditions. The presence and expression of flab, FliS, DnaK, luxs, CsrA, Cj0688, and cosR genes involved in biofilm formation were investigated. Finally, the correlation between the biofilm forming ability of Campylobacter isolates and the presence/expression of selected genes has been explored. A significant correlation was observed between the presence and expression of some genes and the degree of biofilm formation in C. jejuni and C. coli isolates. A strong biofilm production was detected in strains harboring all selected genes with greater expression levels. The ability of C. jejuni and C. coli strains in biofilm formation is associated with the coordinated function and convergent expression of the selected genes. Seemingly, stress response- and motility-related genes have the most involvement in biofilm formation of C. jejuni and C. coli strains, while other genes have an accessory role in this phenomenon.

Immunomodulatory Effects of Adipose-derived Mesenchymal Stem Cells on Epithelial Cells Function in Response to Vibrio cholera in a Co-culture Model.

Inflammation-induced by the interaction of the Vibrio cholerae with the epithelial cells is considered as a main cause of bacteria spreading through the gastrointestinal tract and its consequences. Because of the immunomodulatory and antibacterial properties of adipose-derived mesenchymal stem cells (AD-MSCs), this study aimed to investigate the effect of AD-MSCs on the interaction of the bacterial-epithelial cell. Caco-2 differentiated to intestinal epithelial cells co-cultured with AD-MSCs in a 1:1 ratio of the surface area of six-well plates, for 48 hours. After exposure to Vibrio cholerae, bacterial attachment and internalization were evaluated. Secretions of interleukin (IL) -6, prostaglandin E2 (PGE2), and nitric oxide (NO) were also measured using ELISA, and Griess assay, respectively. In addition, the expression of chloratoxin (Ctx-ß) and inflammatory cytokines such as TNF-α, IL-1ß, and IL-8 were evaluated by real-time polymerase chain reaction (RT-PCR). The rate of apoptosis was also evaluated by Annexin V-PI flow cytometry. Bacterial attachment and Ctx-ß expression were significantly reduced in the co-culture group compared to the Vibrio cholerae-exposed Caco-2. IL-6 and PGE2 secretion increased in the co-culture group. NO, was also slightly reduced in exposure to Vibrio cholerae. An elevated level of bacterial internalization was observed in the co-culture group compared to the Caco-2 cells leading to an increase in the expression of pro-inflammatory cytokines. The rate of apoptosis was also increased significantly. Cell-to-cell contact of AD-MSCs and Caco-2 promoted inflammatory responses and disruption of the epithelium barrier by enhancing bacterial invasion. This may be due to the high expression of surface matrix metalloproteinases on MSCs.

Methicillin-resistant Staphylococcus aureus colonization of infectious and non-infectious skin and soft tissue lesions in patients in Tehran.

BACKGROUND: The most common clinical manifestations of Staphylococcus aureus strains in the community are skin and soft-tissue infections. S. aureus could colonize the body sites and complicate the pathogenesis of skin diseases. S. aureus colonization is a risk factor for severe conditions such as bone and joint infections, pneumonia, bacteremia, and endocarditis. This study aimed to investigate the prevalence of S. aureus strains in skin and soft tissue infections and other skin disorders in patients referring to dermatology clinics and to evaluate the antibiotic resistance pattern and molecular characteristics of S. aureus isolates. METHODS: Skin swabs were collected from the lesional sites in 234 outpatients referring to dermatology clinics in three hospitals in Tehran. Antibiotic susceptibility, biofilm formation, and hemolysis tests were performed for isolates. PCR was done for SCCmec typing, agr grouping, and virulence genes detecting. RESULTS: The prevalence of S. aureus strains among patients with skin and soft-tissue infections and other skin lesions was 44.77% (30/67) and 44.91% (75/167), respectively. Also, 59 (56.19%) isolates were MRSA, 35.57% were HA-MRSA, and 30.5% were CA-MRSA. The psmα gene was more prevalent (62.8%) among isolates, followed by hlaα (56.1%), tsst-1 (15.2%) eta (13.3%), etb (6.6%), and pvl (2.8%). The agr specificity groups I, II, III, and IV were identified in 49.5, 21.9, 11.4, and 14.2% of S. aureus isolates, respectively. Most (56%) S. aureus isolates produced a moderate biofilm, and 23.8% of them produced strong biofilms. α-hemolysin (46.6%), ß-hemolysin (25.7%), γ-hemolysin (19%), and both α and ß-hemolysin (5.7%) were also produced by isolates. CONCLUSION: The present study results indicated high colonization of skin lesions by HA-MRSA and CA-MRSA clones; MRSA strains were more resistant to antibiotics, contained various toxin genes, and were able to form biofilms. Therefore, they could play a vital role in the pathogenesis of various skin diseases; also, they could spread and cause infections in other body sites. Eradication and decolonization strategies could prevent recurrent infections and the spread of resistant strains and improve skin conditions.

Dual-responsive nisin loaded chondroitin sulfate nanogel for treatment of bacterial infection in soft tissues.

Chondroitin sulfate-Nisin nanogels (CS-N NGs) were prepared by electrostatic interaction for nisin delivery as an antibacterial agent in the treatment of bacterial infections caused by some clinical strains of methicillin-resistant and methicillin-sensitive Staphylococcus aureus (S. aureus). The physical properties of CS-N NGs were evaluated using Fourier-transform infrared spectroscopy, dynamic light scattering, and field emission scanning electron microscopy. The average diameter of obtained CS-N NGs was about 65 nm and the stability of nanogels was assessed by zeta potential measurement. Enzyme and pH-responsibility of CS-N NGs due to the presence of susceptible bonds in chondroitin sulfate resulted in effective and controlled release of nisin in the simulated infectious medium. Also, the ability of prepared CS-N NGs for eradicating clinical methicillin resistance S. aureus strain was confirmed by Broth Microdilution Method (BMD) and the cytotoxicity analysis was carried out on Human Dermal Fibroblast (HDF) cells by MTT assay method. Based on the results, this versatile drug carrier could efficiently deliver the cationic antimicrobial peptides as a natural antibiotic for growth inhibition of methicillin-resistant S. aureus strains and further destroying the bacteria in the treatment of subcutaneous infections caused by methicillin-resistant S. aureus strains.

Investigating the effect of an identified mutation within a critical site of PAS domain of WalK protein in a vancomycin-intermediate resistant Staphylococcus aureus by computational approaches.

BACKGROUND: Vancomycin-intermediate resistant Staphylococcus aureus (VISA) is becoming a common cause of nosocomial infections worldwide. VISA isolates are developed by unclear molecular mechanisms via mutations in several genes, including walKR. Although studies have verified some of these mutations, there are a few studies that pay attention to the importance of molecular modelling of mutations. METHOD: For genomic and transcriptomic comparisons in a laboratory-derived VISA strain and its parental strain, Sanger sequencing and reverse transcriptase quantitative PCR (RT-qPCR) methods were used, respectively. After structural protein mapping of the detected mutation, mutation effects were analyzed using molecular computational approaches and crystal structures of related proteins. RESULTS: A mutation WalK-H364R was occurred in a functional zinc ion coordinating residue within the PAS domain in the VISA strain. WalK-H364R was predicted to destabilize protein and decrease WalK interactions with proteins and nucleic acids. The RT-qPCR method showed downregulation of walKR, WalKR-regulated autolysins, and agr locus. CONCLUSION: Overall, WalK-H364R mutation within a critical metal-coordinating site was presumably related to the VISA development. We assume that the WalK-H364R mutation resulted in deleterious effects on protein, which was verified by walKR gene expression changes.. Therefore, molecular modelling provides detailed insight into the molecular mechanism of VISA development, in particular, where allelic replacement experiments are not readily available.