Thiyagarajan Yugendran1, Belgode Narasimha Harish2. 1. Research Scholar, Department of Microbiology, JIPMER , Puducherry, India . 2. Senior Professor and Head, Department of Microbiology, JIPMER , Puducherry, India .
Abstract
INTRODUCTION: Fluoroquinolone resistant clinical isolates belonging to the family Enterobacteriaceae is a major public health concern in India. Data analysis in JIPMER hospital revealed 10% rise in fluoroquinolone resistance within a span of three years suggestive of the possible involvement of mechanism/s other than QRDR capable of imparting fluoroquinolone resistance. DNA methylation regulates gene expression. Moreover, methylated cytosine is a mutational hotspot. Thus, DNA methylation can alter bacterial gene expression profile as well as facilitate the bacteria in accumulating mutations possibly leading to increased antimicrobial resistance. Therefore, the present study was carried out to identify the potential involvement of DNA methylation in ciprofloxacin resistance. AIM: To elucidate and compare the methylation level of genomic and plasmid DNA among clinical isolates of E. coli sensitive and resistant to ciprofloxacin. MATERIALS AND METHODS: The study included 40 clinical E. coli isolates of which, 30 were ciprofloxacin-resistant and 10 were sensitive to ciprofloxacin. Genomic DNA (gDNA) and plasmid DNA were extracted and quantified. Methylation levels were elucidated using 5-mC DNA ELISA kit (Zymoresearch, California, USA) as per kit protocol and guidelines. STATISTICAL ANALYSIS: Spearman correlation 2-tailed test. A p-value <0.05 was considered significant. RESULTS: The MIC values of sensitive and resistant strains against ciprofloxacin ranged from 0.125 μg/mL - 0.75 μg/mL and 8 μg/mL - >256 μg/mL respectively. No difference was found in plasmid DNA methylation level but, the gDNA methylation level of the resistant strains significantly differed from that of the sensitive strains. Based on Spearman correlation test gDNA methylation level of bacteria was found to be inversely proportional to its MIC against ciprofloxacin with p= -0.956 (p-value < 0.0001). CONCLUSION: The influence of DNA methylation over plasmid-mediated quinolone resistance needs to be further confirmed by bisulphite DNA sequencing of the plasmid-borne genes. Extensive usage of ciprofloxacin has led to rise in ciprofloxacin resistance possibly induced by DNA methylation. Thus rational usage of ciprofloxacin in a clinical setting is essential to combat the further development of ciprofloxacin resistance. Hypomethylated genes and adenine methylation needs to identified to fill up gaps in knowledge concerning the involvement of DNA methylation in fluoroquinolone resistance exhibited by E. coli.
INTRODUCTION:Fluoroquinolone resistant clinical isolates belonging to the family Enterobacteriaceae is a major public health concern in India. Data analysis in JIPMER hospital revealed 10% rise in fluoroquinolone resistance within a span of three years suggestive of the possible involvement of mechanism/s other than QRDR capable of imparting fluoroquinolone resistance. DNA methylation regulates gene expression. Moreover, methylated cytosine is a mutational hotspot. Thus, DNA methylation can alter bacterial gene expression profile as well as facilitate the bacteria in accumulating mutations possibly leading to increased antimicrobial resistance. Therefore, the present study was carried out to identify the potential involvement of DNA methylation in ciprofloxacin resistance. AIM: To elucidate and compare the methylation level of genomic and plasmid DNA among clinical isolates of E. coli sensitive and resistant to ciprofloxacin. MATERIALS AND METHODS: The study included 40 clinical E. coli isolates of which, 30 were ciprofloxacin-resistant and 10 were sensitive to ciprofloxacin. Genomic DNA (gDNA) and plasmid DNA were extracted and quantified. Methylation levels were elucidated using 5-mC DNA ELISA kit (Zymoresearch, California, USA) as per kit protocol and guidelines. STATISTICAL ANALYSIS: Spearman correlation 2-tailed test. A p-value <0.05 was considered significant. RESULTS: The MIC values of sensitive and resistant strains against ciprofloxacin ranged from 0.125 μg/mL - 0.75 μg/mL and 8 μg/mL - >256 μg/mL respectively. No difference was found in plasmid DNA methylation level but, the gDNA methylation level of the resistant strains significantly differed from that of the sensitive strains. Based on Spearman correlation test gDNA methylation level of bacteria was found to be inversely proportional to its MIC against ciprofloxacin with p= -0.956 (p-value < 0.0001). CONCLUSION: The influence of DNA methylation over plasmid-mediated quinolone resistance needs to be further confirmed by bisulphite DNA sequencing of the plasmid-borne genes. Extensive usage of ciprofloxacin has led to rise in ciprofloxacin resistance possibly induced by DNA methylation. Thus rational usage of ciprofloxacin in a clinical setting is essential to combat the further development of ciprofloxacin resistance. Hypomethylated genes and adenine methylation needs to identified to fill up gaps in knowledge concerning the involvement of DNA methylation in fluoroquinolone resistance exhibited by E. coli.
Authors: Ari Robicsek; Jacob Strahilevitz; George A Jacoby; Mark Macielag; Darren Abbanat; Chi Hye Park; Karen Bush; David C Hooper Journal: Nat Med Date: 2005-12-20 Impact factor: 53.440
Authors: Christina Kahramanoglou; Ana I Prieto; Supriya Khedkar; Bettina Haase; Ankur Gupta; Vladimir Benes; Gillian M Fraser; Nicholas M Luscombe; Aswin S N Seshasayee Journal: Nat Commun Date: 2012-06-06 Impact factor: 14.919