POC for Quantifying Carbapenem produced KPC from Rectal Swabs in hours

Abstract Carbapenem resistant Enterobacteriaceae CRE pose a serious threat to public health These organisms cause infections that are associated with high mortality rates have the potential to spread widely and are resistant to all lactam agents and most other antimicrobials leaving virtually no treatment options Much CRE increase has been due to the spread of CRE that produce the carbapenemase Klebsiella pneumoniae Carbapenemase KPC enzyme KPC in health care settings is a significant challenge as mortality for patients infected with KPC was percent in days percent in days and percent by the end of their hospitalization To reduce the risk of KPC outbreaks and minimize associated healthcare costs we propose to develop an ultra sensitive point of care POC test for quantifying KPC in about hours from rectal swabs using a POC cartridge The test will confirm that Klebsiella pneumoniae is present viable and capable of producing KPC enzymes when exposed to a carbapenem and it will also indicate the KCE concentration to medical professionals which can provide unique insight for treatment options The proposed KPC guanine amplified immunoassay will quantify ultra low levels of KPC in hours from rectal swabs using a POC cartridge Rectal swab samples will be exposed to a carbapenem antibiotic along with nutrients and incubated for hours at O C to produce KPC if Klebsiella is present viable and carbapenemase producing The test will quantify low levels of KPC using a novel signal amplification platform in a sandwich immunoassay that replaces single optical labels with millions of electroactive guanine rich oligonucleotide tags bound to microbeads Capture antibodies bound to immunomagnetic beads will magnetically separate targets from nonspecific materials Detection limits can be adjusted by changing the size of the microbeads and or the length of the oligonucleotides KPC enzyme levels will be quantified from voltammetry signals using a commercial off the shelf well biosensor micro plate and electrochemical reader potentiostat The feasibility of the Guanine amplified immunoassay is supported by a US Navy project which detected cfu mL of E coli O H and a US Army project for simultaneously detecting E coli cryptosporidium and norovirus Independent testing by demonstrated a log reduced level in detecting C parvum antigens from Guanine s immunoassay compared with commercial ELISA kits In Phase I we will develop a Guanine amplified immunoassay for quantifying low level KPC enzymes produced by CP Klebsiella pneumoniae exposed to carbapenem antibiotics within hours from rectal swab samples Mo adapt the KPC Guanine amplified immunoassay to operate in a cartridge for automated POU Mo and evaluate the KPC Guanine amplified immunoassay versus Modified Hodge Test PCR and culture Mo In Phase expanded testing will be used in applications to the FDA for certification An automated instrument will also be developed to operate the cartridge at the POC for rapid ultra sensitive multiplex quantification of KPC and other CRE Guanine Inc Confidential and Proprietary Page Narrative Carbapenem resistant Enterobacteriaceae CRE cause infections associated with high mortality rates have the potential to spread widely and are resistant to all lactam agents and most other antimicrobials leaving virtually no treatment options Most common CRE tests involve time intensive culturing techniques which can take days to confirm the presence of KCE PCR tests are not widely available leaving healthcare settings vulnerable The proposed project will develop a POC for quantifying KPC from rectal swabs in hours Guanine Inc Confidential and Proprietary Page