شناسایی گونه های استرپتومایسسی مولد آنتی بیوتیک های ضد میکروبی از خاک ایران با روشهای فنوتایپی وژنوتایپی

چکیده : مقدمه: اکتینومیسیت ها باکتری هایی گرم مثبت و رشته ای هستند که بخش اعظم میکروارگانیسم های خاك را در برمی گیرند . بر اساس مطالعات انجام یافته، سه چهارم از کل آنتی بیوتیک های شناخته شده را اکتینو میسیت ها تولید می کنند. در همین راستا غربالگری برای فعالیت ضدباکتریایی ونیز شناسایی آنها از ژن 16S rRNA و روشهای فنوتایپی استفاده شد. هدف این مطالعه شناسایی اکتینومیسیت های مناطق مختلف ایران از جهت خواص آنتی باکتریائی و بررسی سویه های فعال با استفاده از ژن 16S rRNA و روش های فنوتایپی می باشد. روش بررسی: ایزوله های اکتینومیست از نمونه خاك های جمع آوری شده جدا سازی شد، غربالگری اولیه به روش Cross streak method درمحیط کشت اگار وغربالگری ثانویه با روش انتشار دیسک در آگار (Disk Diffusion Method)در مقابل میکروارگانیسم های آزمایشی : ATCC 25923 S. aureusو ATCC 25922 E. coli انجام شدند. تایید نهایی نوع آنتی بیوتیک تولیدی با HPLC و شناسایی سویه های مولد با PCR و تعیین توالی DNA صورت گرفت. یافته ها : از 100 نمونه خاك جمع آوری شده،52 ایزوله اکتینومیست جدا شد،30 ایزوله درغربالگری اولیه و3 ایزوله درغربالگری ثانویه انتخاب شد، سویه 28 دارای پیک (RF) مشابه جنتامیسین و سویه 34و 4 دارای پیک مشابه استرپتومایسین بودند. ژن 16S rRNA ایزوله ها توالی یابی شدند که ایزوله 28 با youssoufiensis Streptomyces 93/99 درصد وایزوله 4 با Streptomyces cyaneofuscatus 93/99 درصد شباهت دارند. نتیجه: نتایج این تحقیق نشان می دهد ایزوله های جدیدی در نمونه خاك های ایران وجود دارد که توانایی تولید مواد آنتی باکتریایی جدیدی دارند. کلمات کلیدی: کلمات کلیدی:استرپتومایسس، ماده ضدباکتریال، جداسازی ازخاك، ژن16S rRN

Synergistic Antiparkinsonian Effect of Flunarizine, Glibenclamide and B Vitamins in a Rate 6-Hydroxydopamine Model; The Role of Malondialdehyde

Background: The current study evaluated the effects of a combination of flunarizine (flu) a calcium channel blocker, glibenclamide (Glib), a KATP channels blocker and B vitamins (B com) on the behavioral symptoms of 6-hydroxydopamine (6-OHDA)-induced model of Parkinson disease to evaluate the synergistic antiparkinsonian effects of the drugs and supplements. Also the level of malondialdehyde (MDA) was measured in blood and brain suspensions to find probable neuroprotective mechanism of these materials. Methods: 6-OHDA was injected into striatum of rats by stereotaxic surgery. Pretreatment with flu, Glib and B com was started before the surgery and continued to three weeks after the surgery. Development and severity of Parkinson disease were evaluated by the conventional behavioral tests. MDA values were measured spectrophotometrically, using thiobarbituric acid test and the MDA standard curve. Results: Pretreatment with a combination of flu, Glib and B com ameliorated the behavioral symptoms of Parkinson disease. The effect of the combination was significantly more potent than those of flu, Glib or B com, solely. Pretreatment with the combination or using only Glib or B com separately, reduced the level of MDA in blood and brain, significantly. However, the effect of the combination was significantly more potent than those of Glib or B com, solely. Conclusions: Since the severity of the behavioral symptoms in the 6-OHDA-induced model of Parkinson disease reflects the degree of the lesion in substantia nigra (SN) dopaminergic neurons, it is suggested that using the combination had neuroprotective effects. The obtained data suggest a synergistic neuroprotective and antiparkinsonian effect for flu, Glib and B com. At least, a part of this effect was mediated through inhibition of oxidative stress. Keywords: 6-Hydroxydopamine, Flunarizine, Glibenclamide: B Vitamins, Behavioral Symptoms, Malondialdehyd

Feasible Relation between Glutathione Peroxidase and Febrile Seizure

How to Cite This Article: Mahyar A, Ayazi P, Dalirani R, Mohammad Hoseini B, Sarookhani MR, Javadi A, Esmaeily Sh. Feasible Relation between Glutathione Peroxidase and Febrile Seizure. Iran J Child Neurol. Winter 2017; 11(1):65-69.AbstractObjectiveWe aimed to determine the relationship between serum glutathione peroxidase and febrile seizure.Materials & MethodsIn this case-control study, 43 children with simple febrile seizure (case group) were compared with 43 febrile children without seizure (control group) in terms of serum glutathione peroxidase level, measured by ELISA method. This study was conducted in Qazvin Children Hospital, Qazvin University of Medical Sciences in Qazvin, Iran in 2012-2013. The results were analyzed and compared in two groups.ResultsFrom 43 children 24 (53%) were male and 19 (47%) were female in children with simple febrile seizure, and 26 (60%) were male and 17 (40%) were female in febrile children without seizure (control group) (P=0.827). Serum glutathione peroxidase level was 166 U/ml (SD=107) in the case group and 141 U/ml (SD=90.5) in the control group of no significant difference.ConclusionThere was no significant relationship between serum glutathione peroxidase and simple febrile seizure. Thus, it seems that glutathione peroxidase, an essential component of antioxidant system, does not play any role in the pathogenesis of simple febrile seizure.References1. Duffner PK, Baumann RJ, Berman P, Green JL, Schneider S, Hodgson ES, etal. Febrile seizures: clinical practice guideline for the long-term management of the child with simple febrile seizures. Pediatrics 2008 ;121:1281-6.Midline doi: 10.1542/peds.2008-0939.2.Shinnar S. Febrile seizures. In: Swaiman KF, Ashwal S,Ferriero DM. Pediatric neurology: principles and practice. 4th ed. Philadelphia: Mosby, 2006:1079-86.3. Mikati M A. Febrile seizures.In: Kliegman RM, Stanton B F, GemeIII J WS, Schor NF , Behrman RE. Nelson textbook of pediatrics.19th ed. Philadelphia: Saunders, 2011:2017-19. 4. Hara K, Tanabe T, Aomatsu T, Inoue N, Tamaki H, Okamoto N ,etal. Febrile seizures associated with influenza A. Brain and Development 2007;29: 30-38.5. Sugai K.Current management of febrile seizures in Japan: An overview . Brain and Development 2010; 32: 64-7. Midline doi: 10.1016/j.braindev.2009.09.019.6. Camfield P, Camfield C, Kurlemann G. Febrile seizures, epileptic syndromes in infancy, childhood, and adolescence. 3th ed. London: John Libbey & Co Ltd,2002:145–52.7. Sapir D, Leitner Y, Harel S, Kraumer U. Unprovoked seizures after complex febrile convulsions. Brain Dev 2000; 22:484–6.8. Kumari PL, Nair MK, Nair SM, Kailas L, Geetha S. Iron deficiency as a risk factor for simple febrile seizures--a case control study. Indian Pediatr 2012; 49:17-9.9. Tutuncuoglu S, Kutukculer N, Kepe L, Coker C, Berdeli A, Tekgul H. Proinflammatory cytokines, prostaglandins and zinc in febrile convulsions. Pediatr Int 2001;43:235-9.10. Ashrafi MR, Shams S, Nouri M, Mohseni M, Shabanian R, Yekaninejad MS, etal. A probable causative factor for an old problem: selenium and glutathione peroxidase appear to play important roles in epilepsy pathogenesis. Epilepsia 2007 ;48:1750-5.11. Willmore IJ, Rubin JJ. Antiperoxidant pretreatment and iron-induced epileptiform discharges in the rat: EEG and histopathologic studies. Neurology1981; 31:63–69. 12.Irshad M, Chaudhuri PS. Oxidant-antioxidant system: role and significance in human body. Indian J Exp Biol 2002 ;40:1233-9.13.Rayman MP.The importance of selenium to human health. Lancet 2000;356:233-41.14.Patel M. Mitochondrial dysfunction and oxidative stress: cause and consequence of epileptic seizures. Free Radical Biology & Medicine 2004;37:1951–1962.15.Li-Ping L, Patel M. Seizure induced changes in mitochondrial redox status. Free Radical Biology & Medicine 2006;40:316–322.16. Weber GF, Maertens P, Meng XZ, Pippenger CE. Glutathione peroxidase deficiency and childhood seizures. Lancet 1991 15; 337:1443-4.17. Sudha K, Rao AV, Rao A. Oxidative stress and antioxidants in epilepsy. Clin Chim Acta 2001; 303:19-24.18. Verrotti A, Basciani F, Trotta D, Pomilio MP, Morgese G, Chiarelli F.Serum Copper, Zinc, Selenium, Glutathione peroxidase and Superoxide dismutase levels in epileptic children before and after 1 year of sodium valproate and carbamazepine therapy. Epilepsy Res 2002;48:71-5.19. Ben-Menachem E. Kyllerman M, Marklund S. Superoxide dismutase and glutathione peroxidase function in progressive myoclonus epilepsies. Epilepsy Res 2000;40:33-9.20. Turkdogan D, Toplan S, Karakoc Y. Lipid peroxidation and antioxidative enzyme activities in childhood epilepsy. J Child Neurol 2002; 17:673-6. 21.Naziroglu M, Kutluhan S, Yilmaz M. Selenium and topiramate modulates brain microsomal oxidative stress values, Ca2+-ATPase activity, and EEG records in pentylenetetrazol-induced seizures in rats. J Membr Biol 2008;225:39-49.22.Naziroglu M. Role of selenium on calcium signaling and oxidative stress-induced molecular pathways in epilepsy. Neurochem Res 2009; 34:2181-91.Medline doi: 10.1007/s11064-009-0015-8.23.Brigelius-Flohé R, Maiorino M.Glutathione peroxidases. Biochim Biophys Acta 2013 ;1830:3289-303. 24. Harapin I, Bauer M, Bedrica L, Potoanjak D. Correlation between gluthathione peroxidase activity and the quantity of selenium in the whole blood of beef calves. Acta Vet Brno 2000; 69: 87–92.Medline. doi: 10.1016/j.bbagen.2012.11.020.25.Koller LD, South PJ, Exon JH, Whitbeck GA, Maas J. Comparison of selenium levels and glutathione peroxidase activity in bovine whole blood. Can J Comp Med 1984 ;48:431-3

Bioinformatics design of CRISPR guide RNA for genomic knockout of ABCB1 gene

Abstract Background: Over-expression of P-Glycoprotein (Pgp) induces acquired drug resistance. Therefore, targeting Pgp as a dominant efflux transporter involved in emergence of multidrug resistance (MDR) has become a major strategy for reversibility of sensitivity to chemotherapy. Objectives: The aim of this study was to design sgRNAs targeting ABCB1 in order to knockout and inhibit the expression of Pgp in Adriamycin resistant (A2780/ADR) ovarian cancer cell line. Methods: This study was performed as a bioinformatics and computational research in Qazvin University of Medical Sciences in collaboration with the Isfahan University of Medical Sciences during 2015-2016. All the 28 exons of the ABCB1 gene were separately investigated in terms of single guide RNA (sgRNA) target sites with regards to the highest on-target and lowest off-target activities, using www.deskgen.com website. Three sgRNA sequences were chosen and synthesized by the GeneCopoeia company. All the plasmids were validated after extraction using BamH1 and EcoR1 restriction enzymes. Results: Sequences of the three sgRNAs were selected close to the start codon (ATG) in order to maximize the possibility of exons 4 and 5 knockout. Digested pCRISPR-CG01, using BamH1 and EcoR1, was electrophorized on 1.5% agarose gel. Detection of the two 330bp and 10100bp fragments on the gel confirmed the integrity of the plasmid and success of the restriction enzyme digestion. Conclusions: The vectors containing the designed sgRNA sequences and CRISPR associated protein (Cas9) can inhibit Pgp gene expression in cell lines over-expressing this gene, including A2780/ADR. Keywords: Drug Resistance, P-Glycoprotein, Ovarian Cancer, CRISP

Synergistic Antiparkinsonian Effect of Flunarizine, Glibenclamide and B Vitamins in a Rat 6-Hydroxydopamine Model; The Role of Malondialdehyde

Background: The current study evaluated the effects of a combination of flunarizine (flu) a calcium channel blocker, glibenclamide (Glib), a KATP channels blocker and B vitamins (B com) on the behavioral symptoms of 6-hydroxydopamine (6-OHDA)-induced model of Parkinson disease to examine the synergistic antiparkinsonian effects of the drugs and supplements. Also the level of malondialdehyde (MDA) was measured in blood and brain suspensions to find probable neuroprotective mechanism of these materials. Methods: 6-OHDA was injected into striatum of rats by stereotaxic surgery. Pretreatment with flu, Glib and B com was started before the surgery and continued to three weeks after the surgery. Development and severity of Parkinson disease were evaluated by the conventional behavioral tests. MDA values were measured spectrophotometrically, using thiobarbituric acid test and the MDA standard curve. Results: Pretreatment with a combination of flu, Glib and B com ameliorated the behavioral symptoms of Parkinson disease. The effect of the combination was significantly more potent than those of flu, Glib or B com, solely. Pretreatment with the combination or using only Glib or B com separately, reduced the level of MDA in blood and brain, significantly. However, the effect of the combination was significantly more potent than those of Glib or B com, solely. Conclusions: Since the severity of the behavioral symptoms in the 6-OHDA-induced model of Parkinson disease reflects the degree of the lesion in substantia nigra (SN) dopaminergic neurons, it is suggested that using the combination had neuroprotective effects. The obtained data suggest a synergistic neuroprotective and antiparkinsonian effect for flu, Glib and B com. At least, a part of this effect was mediated through inhibition of oxidative stress. Keywords: 6-Hydroxydopamine, Flunarizine, Glibenclamide: B Vitamins, Behavioral Symptoms, Malondialdehyd

Detection of Extended-Spectrum beta-Lactamases among Acinetobacter Baumannii Isolated from Hospitals of Qazvin, Iran

BACKGROUND፡ Acinetobacter baumannii is a major contributor to nosocomial infections. Extended-spectrum ßlactamase (ESBL)-producing A. baumannii is spreading worldwide. We aimed to determine the frequency of ESBLencoding genes in clinical isolates of A. baumannii and to access their clonal relationship by repetitive extragenic palindromicPCR (rep-PCR). METHODS: In this descriptive cross-sectional study, 203 isolates of A. baumannii were collected from Qazvin hospitals. The Identification of isolates was performed by standard laboratory methods. To verify ESBL production, all isolates were screened by disk agar diffusion and confirmed by the combined disk method. Subsequently, ESBL-encoding genes were detected by PCR and sequencing. Possible clonal association of ESBL-producing isolates was evaluated using rep-PCR. RESULTS: Two hundred (98.5%) isolates showed reduced susceptibility to one of the antibiotics used in the ESBL screening test, of which 127 isolates (62.6%) produced ESBL. PCR results showed blaOXA-1 (20.5%) was the most prevalent gene followed by blaTEM-1 (20%), blaGES-1 (15.7%), blaCTX-M-15 (7.9%), and blaPER-1 (1.6%). Rep-PCR results revealed that ESBL-producing isolates belonged to clones A (85%), B (13.4%), and C (1.6%). CONCLUSION: Our study showed the significant presence of blaOXA-1, blaTEM-1, blaGES-1, blaCTX-M-15, and blaPER-1 genes in ESBLproducing A. baumannii isolates in the studied hospitals. This is the first report on the emergence of blaOXA-1 gene in these isolates in Iran. The use of comprehensive antimicrobial treatment guidelines based on laboratory data and appropriate infection control interventions are essential