02451nas a2200193   4500008004100000245025800041210006900299260000800368490000700376520146600383653013101849100002501980700002602005700002702031700002502058700003102083700002102114856012202135       2017                        eng d00aScalable metabolite supplementation strategy against antibiotic resistant pathogen Chromobacterium violaceum induced by NAD(+)/NADH(+) imbalanceA scalable metabolite supplementation strategy against antibiotic resistant pathogen Chromobacterium violaceu0 aScalable metabolite supplementation strategy against antibiotic   cAPR0 v113 a<p>BACKGROUND: The leading edge of the global problem of antibiotic resistance necessitates novel therapeutic strategies. This study develops a novel systems biology driven approach for killing antibiotic resistant pathogens using benign metabolites. RESULTS: Controlled laboratory evolutions established chloramphenicol and streptomycin resistant pathogens of Chromobacterium. These resistant pathogens showed higher growth rates and required higher lethal doses of antibiotic. Growth and viability testing identified malate, maleate, succinate, pyruvate and oxoadipate as resensitising agents for antibiotic therapy. Resistant genes were catalogued through whole genome sequencing. Intracellular metabolomic profiling identified violacein as a potential biomarker for resistance. The temporal variance of metabolites captured the linearized dynamics around the steady state and correlated to growth rate. A constraints-based flux balance model of the core metabolism was used to predict the metabolic basis of antibiotic susceptibility and resistance. CONCLUSIONS: The model predicts electron imbalance and skewed NAD/NADH ratios as a result of antibiotics - chloramphenicol and streptomycin. The resistant pathogen rewired its metabolic networks to compensate for disruption of redox homeostasis. We foresee the utility of such scalable workflows in identifying metabolites for clinical isolates as inevitable solutions to mitigate antibiotic resistance.</p>10aAntibiotic resistance; Flux balance analysis; Flux variability analysis; Metabolism; Metabolomic; NAD; NADH; Redox homeostasis1 aBanerjee, Deepanwita1 aParmar, Dharmeshkumar1 aBhattacharya, Nivedita1 aGhanate, Avinash, D.1 aPanchagnula, Venkateswarlu1 aRaghunathan, Anu  uhttps://library.ncl.res.in/content/scalable-metabolite-supplementation-strategy-against-antibiotic-resistant-pathogen01851nas a2200169   4500008004100000245009800041210006900139260000800208300001000216490000800226520121500234100003201449700002501481700003101506700002101537856012301558       2018                        eng d00a Integrated constraints based analysis of an engineered violacein pathway in escherichia coli0 aIntegrated constraints based analysis of an engineered violacein  cSEP  a10-190 v1713 aStrategies towards optimal violacein biosynthesis, a potential drug molecule, need systems level coordination of enzymatic activities of individual genes in a multigene operon vioABCDE. Constraints-based flux balance analysis of an extended iAF1260 model (iAF1260vio) with a reconstructed violacein module predicted growth and violacein yields in Escherichia coli accurately. Shadow price (SP) analysis identified tryptophan metabolism and NADPH as limiting. Increased tryptophan levels in Delta pgi & Delta pheA were validated using in silico gene deletion analysis. Phenotypic phase plane (PhPP) analysis highlighted sensitivity between tryptophan and NADPH for violacein synthesis at molar growth yields. A synthetic VioABCDE operon (SYNO) sequence was designed to maximize Codon Adaptive Index (CAI: 0.9) and tune translation initiation rates (TIR: 2-50 fold higher) in E. coli. All pSYN E. coli transformants produced higher violacein, with a maximum six-fold increase in yields. The rational design E. coli: Delta pheA SYN: gave the highest violacein titers (33.8 mg/I). Such integrated approaches targeting multiple molecular hierarchies in the cell can be extended further to increase violacein yields.1 aImmanuel, Selva, Rupa Chris1 aBanerjee, Deepanwita1 aRajankar, Mayooreshwar, P.1 aRaghunathan, Anu  uhttps://library.ncl.res.in/content/integrated-constraints-based-analysis-engineered-violacein-pathway-escherichia-coli01307nas a2200145   4500008004100000245012400041210006900165260000800234300001400242490000800256520072100264100002500985700002101010856013001031       2018                        eng d00aKnowledge, attitude and practice of antibiotic use and antimicrobial resistance: a study post the 'Red Line' initiative0 aKnowledge attitude and practice of antibiotic use and antimicrob  cMAY  a1866-18770 v1143 aAntimicrobial resistance (AMR) is propagated by irrational use of antibiotics by healthcare practitioners and the uninformed public. This study assessed a select cohort of 504 urban Indians for their knowledge, awareness and practice of antibiotic use and AMR. Forty seven per cent were unaware of the differences between over-the-counter drugs and antibiotics. One in four believes that dose-skipping does not contribute to AMR. One in ten tends to self-medicate. One in five bought medicines without prescription or started an antibiotic course by calling a doctor. Our results mandate educational campaigns, stewardship and surveillance at the national level for prudent antimicrobial use in the Indian community.1 aBanerjee, Deepanwita1 aRaghunathan, Anu  uhttps://library.ncl.res.in/content/knowledge-attitude-and-practice-antibiotic-use-and-antimicrobial-resistance-study-post-red02546nas a2200157   4500008004100000022001400041245014100055210006900196260000900265300002900274490000700303520190900310100002502219700002102244856012302265       2019                        eng d  a1932-620300aConstraints-based analysis identifies NAD(+) recycling through metabolic reprogramming in antibiotic resistant Chromobacterium violaceum0 aConstraintsbased analysis identifies NAD recycling through metab  cJAN   aArticle Number: e02100080 v143 a<p>In the post genomic era, high throughput data augment stoichiometric flux balance models to compute accurate metabolic flux states, growth and energy phenotypes. Investigating altered metabolism in the context of evolved resistant genotypes potentially provide simple strategies to overcome drug resistance and induce susceptibility to existing antibiotics. A genome-scale metabolic model (GSMM) for Chromobacterium violaceum, an opportunistic human pathogen, was reconstructed using legacy data. Experimental constraints were used to represent antibiotic susceptible and resistant populations. Model predictions were validated using growth and respiration data successfully. Differential flux distribution and metabolic reprogramming were identified as a response to antibiotics, chloramphenicol and streptomycin. Streptomycin resistant populations (StrpR) redirected tricarboxylic acid (TCA) cycle flux through the glyoxylate shunt. Chloramphenicol resistant populations (ChlR) resorted to overflow metabolism producing acetate and formate. This switch to fermentative metabolism is potentially through excess reducing equivalents and increased NADH/NAD ratios. Reduced proton gradients and changed Proton Motive Force (PMF) induced by antibiotics were also predicted and verified experimentally using flow cytometry based membrane potential measurements. Pareto analysis of NADH and ATP maintenance showed the decoupling of electron transfer and ATP synthesis in StrpR. Redox homeostasis and NAD(+) cycling through rewiring metabolic flux was implicated in re-sensitizing antibiotic resistant C. violaceum. These approaches can be used to probe metabolic vulnerabilities of resistant pathogens. On the verge of a post-antibiotic era, we foresee a critical need for systems level understanding of pathogens and host interaction to extend shelf life of antibiotics and strategize novel therapies.</p>1 aBanerjee, Deepanwita1 aRaghunathan, Anu  uhttps://library.ncl.res.in/content/constraints-based-analysis-identifies-nad-recycling-through-metabolic-reprogramming