A RAPID ASSAY FOR DETECTION OF DETECTION OF CARBAPENEMASE PRODUCING ORGANISMS

DESCRIPTION provided by applicant A recent outbreak of carbapenem resistant Enterobacteriaceae CRE associated with contaminated duodenoscopes at the Ronald Reagan UCLA Medical Center highlights the importance of early detection of these clinically significant pathogens Coincidentally the Obama Administration released The National Action Plan to Combat Antibiotic Resistant Bacteria for which one of the steps of the action plan is to slow the emergence of resistant bacteria and prevent the spread of resistant infections Detection and identification of carbapenemase producing organisms CPO in health care centers is the first step to confining the source and preventing the potential spread of these multidrug resistant pathogens However detection of CPO in clinical laboratories is challenging as isolates may only have moderate reductions in susceptibilities to carbapenems and resistance may be mediated by other mechanisms i e chromosomal and plasmid mediated cephalosporinases or extended spectrum lactamase ESBL producers with decreased membrane permeability Molecular methods for the detection of carbapenemase genes are limited by the number of carbapenemase targets assayed in addition to other limitations The reference phenotypic method for detection of carbapenemase producing Enterobacteriaceae the Modified Hodge test suffers from a long turn around time lacks specificity and has poor sensitivity for certain types of carbapenemase enzymes i e metallo lactamase enzymes This project aims to develop a low cost easy to use essentially one manual step and rapid min assay to identify carbapenemase production in carbapenem resistant Gram negative bacteria suitable for surveillance and clinical laboratory use The proposed assay uses a luciferin derivatized lactamase substrate for detection of lactamase activity All reagents will be formulated in a master mix Reagent I such that the assay involves essentially one manual step addition of the sample to the master mix To differentiate carbapenemases from other lactamases a second master mix containing a carbapenem will be formulated Reagent II The signal of Reagent I indicates the presence or absence of lactamase while the signal ratio of Reagent II to Reagent I indicates whether the lactamase is a carbapenemase The proposed Phase I project will optimize the assay and use well characterized isolates as well as prospective clinical isolates to evaluate the assay Successful completion of this Phase I project will pave the way for a Phase II study which will involve multiple sites and substantially larger sample numbers A simple and rapid assay for detection of CPO will play a significant role in surveillance and help guide therapy and infection control measures PUBLIC HEALTH RELEVANCE A recent outbreak of carbapenem resistant Enterobacteriaceae CRE associated with contaminated duodenoscopes at the Ronald Reagan UCLA Medical Center highlights the importance of early detection of these clinically significant pathogens Detection and identification of carbapenemase producing organisms CPO in health care centers is the first step to confining the source and preventing the potential spread of these multidrug resistant pathogens The proposed project to develop a simple and rapid assay for detection of CPO if successful will play a significant role in surveillance and help guide therapy and infection control measures