Phenotypic and genotypic evaluation of fluoroquinolone resistance in clinical isolates of Staphylococcus aureus in Tehran

Background: Fluoroquinolones are broad-spectrum antibiotics widely used in the treatment of bacterial infections such as Staphylococcus aureus isolates. Resistance to these antibiotics is increasing. Material/Methods: The occurrence of mutations in the grlA and gyrA loci were evaluated in 69 fluoroquinolone-resistant S. aureus isolates from 2 teaching hospitals of Tehran University of Medical Sciences. Results: Out of the 165 S. aureus isolates, 87 (52.7) were resistant to methicillin and 69 (41.8) were resistant to fluoroquinolone. Fluoroquinolone-resistant S. atoms isolates had a mutation at codon 80 in the grlA gene and different mutational combinations in the gyrA gene. These mutational combinations included 45 isolates at codons 84 and 86,23 isolates at codons 84,86 and 106 and 1 isolate at codons 84, 86 and 90. Fluoroquinolone-resistant S. aureus isolates were clustered into 33 PFGE types. Conclusions: The findings of this study show that the fluoroquinolone-resistant S. aureus strains isolated in the teaching hospitals in Tehran had multiple mutations in the QRDRs region of both grlA and gyrA genes

Regulation of connexin 43 and microRNA expression via β2-adrenoceptor signaling in 1321N1 astrocytoma cells

Connexin 43 (Cx43) is the main gap junction protein in astrocytes and exerts the same effects on growth inhibition in astrocytoma and glioma as microRNA-146a (miR-146a) in glioma. β2-adrenergic receptor (AR) signaling modulates Cx43 expression in myocytes via components downstream of protein kinase A (PKA) and exchange protein directly activated by cAMP (Epac). However, it remains to be elucidated how expression of Cx43 is modulated in astrocytes. In the present study, 1321N1 astrocytoma cells were treated with β2-AR signaling agents in order to evaluate the expression of Cx43 and miRNAs. RNA and protein were extracted from the cells for use in reverse transcription-quantitative polymerase chain reaction and western blot analysis, respectively. The results revealed that clenbuterol increased miR-146a level and upregulated Cx43 expression via cAMP/PKA at the mRNA and protein level. Pre-inhibition of adenyl cyclase decreased expression of Cx43 and miR-146a. PKA activation and overexpression of miR-146a in A-1321N1 cells increased the expression of Cx43. β2-AR stimulation and 6Bnz, a PKA activator, suppressed oncomiRs miR-155 and miR-27a, while 8-(4-chlorophenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphate, an Epac activator, increased their levels. The current findings demonstrated that β2-AR signaling has growth inhibitory effects via modulation of the cAMP/PKA pathway in A-1321N1 cells through increasing the expression level of Cx43 and miR-146a as well as decreasing miR-155 and miR-27a levels. Thus, stimulation of the β2-AR and PKA signaling pathway may be a useful approach for astrocytoma therapy

Combinations of β-lactam or aminoglycoside antibiotics with plectasin are synergistic against methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

Bacterial infections remain the leading killer worldwide which is worsened by the continuous emergence of antibiotic resistance. In particular, methicillin-sensitive (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA) are prevalent and the latter can be difficult to treat. The traditional strategy of novel therapeutic drug development inevitably leads to emergence of resistant strains, rendering the new drugs ineffective. Therefore, rejuvenating the therapeutic potentials of existing antibiotics offers an attractive novel strategy. Plectasin, a defensin antimicrobial peptide, potentiates the activities of other antibiotics such as β-lactams, aminoglycosides and glycopeptides against MSSA and MRSA. We performed in vitro and in vivo investigations to test against genetically diverse clinical isolates of MSSA (n = 101) and MRSA (n = 115). Minimum inhibitory concentrations (MIC) were determined by the broth microdilution method. The effects of combining plectasin with β-lactams, aminoglycosides and glycopeptides were examined using the chequerboard method and time kill curves. A murine neutropenic thigh model and a murine peritoneal infection model were used to test the effect of combination in vivo. Determined by factional inhibitory concentration index (FICI), plectasin in combination with aminoglycosides (gentamicin, neomycin or amikacin) displayed synergistic effects in 76-78% of MSSA and MRSA. A similar synergistic response was observed when plectasin was combined with β-lactams (penicillin, amoxicillin or flucloxacillin) in 87-89% of MSSA and MRSA. Interestingly, no such interaction was observed when plectasin was paired with vancomycin. Time kill analysis also demonstrated significant synergistic activities when plectasin was combined with amoxicillin, gentamicin or neomycin. In the murine models, plectasin at doses as low as 8 mg/kg augmented the activities of amoxicillin and gentamicin in successful treatment of MSSA and MRSA infections. We demonstrated that plectasin strongly rejuvenates the therapeutic potencies of existing antibiotics in vitro and in vivo. This is a novel strategy that can have major clinical implications in our fight against bacterial infections

A study on the chemical characteristics changes throughout the manufacture and ripening of Lighvan cheese

Lighvan cheese is one of the traditional cheeses which have the most high quantity of use in Iran. It is produced in South East of Tabriz in North West of Iran. The raw milk of ewe together with 20% -30% of goat's milk, without yeast, are used for its production. Its taste is mild salty and its scent is pleasant. The purpose of this study was to investigate the chemical indexes changes including salt percentage, the degree of acidity, pH, dry mater, ashes, and protein during the production and ripening. For this purpose, after coordinating with 10 local cheese producers, one batch from each producer and from each batch 20 tins, weighing 1 kg, which in total was 200 newly packaged cheese were purchased randomly. The tins were kept in special caves for 30 days in the region and then 60 days in refrigerator. In each batch sample of the raw milk, clot after rising and before salting, the cheese during the package time in tin and the cheese sample on 15th, 30th, 60th, and 90th days of ripining was analyzed chemically. The results of the study from the initial days of production to the end of the ripening period indicated the following changes: the rate of fat from 6.8 ± 0.25 in milk to 24.55±0.95 in samples, pH from 5.94± 0.06 in milk to 4.4±0.11 in samples, acidity from 39.4 ± 5.99 D° in milk to 119.4±5.38 in samples, rate of ash from 1.77±0.23 in milk to 8.09±2.32 in samples, the percentage of dry mater from 16.52±0.74 in milk to 43.57±1.34 in samples, and finally the percentage of protein from 4.45±1/12 in milk to 14.2±1.4 in samples. This result suggests that Lighvan cheese has unique characteristics in terms of its alteration procedure and chemical characteristics and based on the standard criterion in Iran, 2344-1, it is white cheese ripened brine in terms of chemical characteristics and fatty cheese in terms of the percentage of fat

Molecular analysis of typical and atypical enteropathogenic Escherichia coli (EPEC) isolated from children with diarrhoea

Diarrhoea continues to be one of the most common causes of morbidity and mortality among infants and children in developing countries. To investigate the incidence, antimicrobial resistance and genetic relationships of enteropathogenic Escherichia coli (EPEC) in children with diarrhoea, a total of 612 stool specimens were collected in Tehran, Iran, and cultured to isolate strains of EPEC. The disc diffusion method was used to determine the susceptibility of the isolates according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. The presence of eae, stx and bfp-A genes was determined by PCR. The genetic relationships between EPEC isolates were determined by pulsed-field gel electrophoresis (PFGE). Out of the 412 strains of E. coli obtained from 612 diarrhoeal stool specimens, 23 (5.6) were identified as EPEC, of which seven (30.4) were classified as typical strains of EPEC and 16 (69.6) were classified as atypical. Out of the 23 EPEC isolates, 69.5 were resistant to ampicillin, 39.1 were resistant to tetracycline and cotrimoxazole, 30.4 were resistant to cefpodoxime, ceftazidime, ceftriaxone and aztreonam, and 26.1 were resistant to imipenem. The isolates were classified into 21 pulsotypes by PFGE profiles. The present study shows that typical and atypical EPEC isolates displayed considerable heterogeneity in PFGE profiles and EPEC infections were only sporadic in Tehran. Overall 69 of isolates were resistant to at least one of the antibiotics tested

Chimeric Self-assembling Nanofiber Containing Bone Marrow Homing Peptide�s Motif Induces Motor Neuron Recovery in Animal Model of Chronic Spinal Cord Injury; an In Vitro and In Vivo Investigation

To date, spinal cord injury (SCI) has remained an incurable disaster. The use of self-assembling peptide nanofiber containing bioactive motifs such as bone marrow homing peptide (BMHP1) as an injectable scaffold in spinal cord regeneration has been suggested. Human endometrial-derived stromal cells (hEnSCs) have been approved by the FDA for clinical application. In this regard, we were interested in investigating the role of BMHP1 in hEnSCs� neural differentiation in vitro and evaluating the supportive effects of this scaffold in rat model of chronic SCI. 1,1-Diphenyl-2-picryl-hydrazyl (DPPH), lactate dehydrogenase (LDH) release, 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay, real-time PCR, and immunocyotochemistry (ICC) were performed as a biocompatibility and neural differentiation evaluations on neuron-like hEnSC-derived cells encapsulated into nanofiber. Nanofiber was implanted into rats and followed by behavioral test, Nissl, luxol fast blue (LFB) staining and immunohistostaining (IHC). Results indicated that cell membrane of neuroblastoma cells were more sensitive than hEnSCs to concentration of proton and cell proliferation decreased with increase of concentration. This effect might be related to oxygen tension and elastic modules of scaffold. -BMHP1 nanofiber induced neural differentiation in hEnSC and decreased GFAP gene and protein as a marker of reactive astrocytes in vitro and in vivo. A reason for this finding might be related to the role of spacer number in induction of mechano-transduction signals. The presented study revealed the chimeric BMHP1 nanofiber induced higher axon regeneration and myelniation around the cavity and motor neuron function was encouraged to improve with less inflammatory response following SCI in rats. These effects were possibly due to nanostructured topography and mechano-transduction signals derived from hydrogel at low concentration. © 2015, Springer Science+Business Media New York

Self-Assembling Peptide Nanofiber Containing Long Motif of Laminin Induces Neural Differentiation, Tubulin Polymerization, and Neurogenesis: In Vitro, Ex Vivo, and In Vivo Studies

Spinal cord injury (SCI) in humans stayed a ruining and healless disorder. Since longer laminin motif (CQAASIKVAV (CQIK)) better mimics conformation of native region in active site than isoleucine-lysine-valine-alanine-valine (IKVAV) and resulted in improved cellular response so, for the first time in this study, CQIK bounded with two glycines spacer and (RADA)4 as a self-assembling peptide nanofiber backbone (-CQIK) was used. The purpose of this study was to investigate the role of -CQIK in neural differentiation of human endometrial-derived stromal cells (hEnSCs) in vitro, tubulin polymerization ex vivo, and assess the supportive effect of this hydrogel in an animal model of chronic SCI. Results disclosed that proton concentration has direct effect on hEnSCs membrane damage but not on neuroblastoma cells. However, cell viability of neuroblastoma encapsulated into -CQIK was higher than hEnSCs at the concentration of 0.125 v/w. Gene expression data confirmed neurogenesis, TH over-expression, and glial fibrillary acidic protein (GFAP) suppression eventually through α6 and β1 integrin site. However, it revealed higher neurogenesis as compared to bone morrow homing peptides (BMHP). Although, Basso, Beattie, Bresnahan (BBB) score of chronic model of SCI in rat was higher than control and phosphate-buffered saline (PBS) group but significantly was less than BMHP group. However, -CQIK had induced neurite outgrowth and myelination and inhibited astrogliosis. Tubulin polymerization data using UV spectroscopy showed higher degree of polymerization. However, tubulin polymerization was dependent on nanofiber concentration. Based on our results, it might be concluded that peptidic nanofiber containing long motif of laminin holds great promise for spinal cord injury recovery with increment of neurogenesis and astrogliosis decrement. © 2015, Springer Science+Business Media New York

Characterisation of genes encoding aminoglycoside-modifying enzymes among meticillin-resistant Staphylococcus aureus isolated from two hospitals in Tehran, Iran

Meticillin-resistant Staphylococcus aureus (MRSA) is a major hospital pathogen and typically shows resistance to multiple antimicrobial agents. The susceptibility patterns of 109 MRSA isolates to aminoglycoside antibiotics were determined by the disk diffusion method. Genes encoding the aminoglycoside-modifying enzymes (AMEs) were targeted by polymerase chain reaction (PCR) assays. All isolates were also subjected to multiplex PCR to determine the distribution of staphylococcal cassette chromosome mec (SCCmec) types in the study population. The rates of resistance to various antibiotics were as follows: kanamycin, 97; tobramycin, 96; gentamicin, 87; amikacin, 93; and netilmicin, 80. The most prevalent AME genes were aac(6')-Ie-aph(2 '') (83) followed by aph(3')-IIIa (71). Coexistence of three AME genes was detected in 21 of isolates. The ant(4')-Ia gene was the least frequent AME gene among MRSA isolates (26). Of the 109 isolates, 106 (97) were identified as SCCmec type III or IIIA and 3 (3) as SCCmec type IV. The majority of MRSA isolates belonged to SCCmec III or IIIA and carried the aac(6')- Ie-aph(2 '') gene, which is consistent with results of susceptibility testing of these isolates against aminoglycosides. (C) 2008 Elsevier B. V. and the International Society of Chemotherapy. All rights reserved

Thermogel nanofiber induces human endometrial-derived stromal cells to neural differentiation: In vitro and in vivo studies in rat

Spinal cord injury (SCI) in humans remains a devastating and incurable disorder. The use of Matrigel, a hydrogelmimicking extracellular matrix, has been suggested as a scaffold for spinal cord regeneration. Human endometrial-derived stromal cells (hEnSCs) are abundant and available in adult stem cells with low immunological incompatibility, which could be considered for cell replacement therapy. The purpose of this study was to investigate the role ofMatrigel in neural differentiation of hEnSCs in vitro and assess the supportive effects of this hydrogel in an animal model of SCI. hEnSCs were isolated and encapsulated into nanofibrous thermogel and cell viability and cell membrane damage were assessed. Encapsulated hEnSCs into Matrigel were treated with neural differentiation medium for 21 days, and then neural genes and protein markers were analyzed using real time-PCR and immunocytochemistry. Matrigel was implanted into rats with SCI and followed for 42 days using a behavioral test. Our study revealed a higher cell viability and neural differentiation in the level of genes and proteins as well as lower cell membrane damage. Substantial recoveries of motor function were observed in animals receiving the Matrigel treatment. The treatment with Matrigel, nanofibrous scaffold, produced beneficial effects on functional recovery following SCI in rats, possibly via assimilation to cytoskeleton fiber, high surface/volume ratio, spatial interconnectivity and containing some adhesive molecules and growth factors, enhancement of antiinflammation, anti-astrogliosis, neuronal extension, and neuronal regeneration effects. © 2014 Wiley Periodicals, Inc