Transcriptional analysis of bla NDM-1 and copy number alteration under carbapenem stress

Abstract

BackgroundNew Delhi metallo beta-lactamase is known to compromise carbapenem therapy and leading to treatment failure. However, their response to carbapenem stress is not clearly known. Here, we have investigated the transcriptional response of bla NDM-1 and plasmid copy number alteration under carbapenem exposure.MethodsThree bla NDM-1 harboring plasmids representing three incompatibility types (IncFIC, IncA/C and IncK) were inoculated in LB broth with and without imipenem, meropenem and ertapenem. After each 1 h total RNA was isolated, immediately reverse transcribed into cDNA and quantitative real time PCR was used for transcriptional expression of bla NDM-1. Horizontal transferability and stability of the plasmids encoding bla NDM-1 were also determined. Changes in copy number of bla NDM-1 harboring plasmids under the exposure of different carbapenems were determined by real time PCR. Clonal relatedness among the isolates was determined by pulsed field gel electrophoresis.ResultsUnder carbapenem stress over an interval of time there was a sharp variation in the transcriptional expression of bla NDM-1 although it did not follow a specific pattern. All bla NDM-1 carrying plasmids were transferable by conjugation. These plasmids were highly stable and complete loss was observed between 92nd to 96th serial passages when antibiotic pressure was withdrawn. High copy number of bla NDM-1 was found for IncF type plasmids compared to the other replicon types.ConclusionThis study suggests that the single dose of carbapenem pressure does not significantly influence the expression of bla NDM-1 and also focus on the stability of this gene as well as the change in copy number with respect to the incompatible type of plasmid harboring resistance determinant.Electronic supplementary materialThe online version of this article (doi:10.1186/s13756-017-0183-2) contains supplementary material, which is available to authorized users.

Hospital Wastewater Releases of Carbapenem-Resistance Pathogens and Genes in Urban India

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IncC helper dependent plasmid-like replication of Salmonella Genomic Island 1

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CXC Chemokines Exhibit Bactericidal Activity against Multidrug-Resistant Gram-Negative Pathogens

“…1A ). NDM-1 presents a considerable challenge to antimicrobial therapy because of its high enzymatic and genetic stability ( 9 , 10 ) and exceptional horizontal mobility among Gram-negative bacterial species ( 11 ). On the basis of prior investigations ( 12 ), CRE were treated with 48 µg/ml CXCL10.…”