Presentation on theme: "Detection of Carbapenemases From a Technologist’s Perspective"— Presentation transcript:
1 Detection of Carbapenemases From a Technologist’s Perspective Courtney Fraser MLS (ASCP)Microbiology Supervisor ACL Laboratories
2 DisclosureI will discuss specific brand names in this presentation. I do not have stock in, nor receive honoraria for any commercial product mentioned in this presentation.
3 IntroductionAutomation is not available for carbapenemase confirmatory testing.Resistance mechanisms are not typically discussed in the classroom for most MLS students.Seasoned Techs currently in the field are not used to resistance in organisms and are unfamiliar with the mechanisms related to resistance.Education needed on antibiotic classifications.
4 Purpose of Presentation Define types of carbapenemases and understand classificationsWhen to testHow to testHow to incorporate instrumentation tools
5 What is a Carbapenemase? First, need to understand what are the carbapenems:ErtapenemImipenemMeropenemDoripenemSecond, the suffix –ase indicates an enzymeA carbapenemase is an enzyme produced by bacteria that hydrolyze (deactivate) carbapenems
7 Carbapenems Large groups MeropenemAll share a common β-lactam ring, β-lactamasestarget the β-lactam ring)differences are in ring stabilization from hydrolysisprotection of ring from β-lactamase by addition oflarge groups blocking the substrate siteImipenemErtapenemLarge groups
8 Why Important?Studies have shown that rapid detection of carbapenemases have attributed to positive patient treatment outcomes.Infection Control emergency – These mechanisms can be plasma-mediated allowing for easy transmission to other organisms.Detection of these resistance mechanisms should initiate contact precautions for patients , thus reducing the spread of nosocomial related infections.Deter inappropriate antibiotic use thus prolonging the efficacy of available antibiotics
10 ACL Laboratories Overview ACL Laboratories is defined as an operating management agreementbetween Advocate Health Care and Aurora Health Care.Two Central Labs (Rosemont, IL., West Allis, WI.)Total of 27 hospitalsRosemont
13 Types of β-Lactamases/Carbapenemases Type A (Ambler) – KPC – 1st described 1998, endemic in NYC since Moving west. 1st isolates in Chicago area in 2/08Type B – Metallo ß-lactamasesFound in Stenotrophomonas maltophiliaEnterobacteriaceae and P. aeruginosa - Reported sporadically in U.S.Watch out for NDM-1 which has spread to Europe (Sweden and England) from IndiaType D – Oxa-40 – Endemic in Acinetobacter baumannii in the Chicago area since 2002Hyper AmpC producersMutations in AmpC promoter and attenuator/promoter regionsPoorly inhibited by lactam—lactamase inhibitor combinationsE. coli produces AmpC (poorly) and is unlike other bacteria with AmpC the gene is not inducibleNo ampR regulatory geneAll the above can give a positive Hodge Test!1313
14 Types of Carbapenemases Enzyme TypeAmbler ClassActivity SpectrumOrganism(s)KPC (1-10)(plasmid)AAll β-lactamsEnterobacteriaceae Ps. aeruginosaSMECarbapenems and aztreonam, but not 3rd/4th Gen cephalosporinsS. marcescens , not plasmidAssociated.NMC–A, IMIMNC = Not metallo carbapenemaseIMI = IMI hydrolyzingΒ-lactamaseSame as for SMEEnterobacter spp.GESGES= Guiana extended spectrum (plasmid)Imipenem and 3rd/4th cephalosporinsPs. Aeruginosa and EnterobacteriaceaeIMI, VIM, NDM-1VIM = Verona Integron encoded MBL) NDM-1 = New Delhi metallo β lactamaseB (metallo-β-lactamases)All β-lactams; can test susceptible to aztreonam(NDM-1 variable AZT resistance)Pseudomonas spp. Acinetobacter spp. EnterobacteriaceaeOXA(Oxacillin hydrolyzing)DWeakly active against carbapenemsA. baumanii, P. Aeruginosa, and rare EnterobacteriaceaeS. marcescensenzymePediatr Infect Dis J. 2010;29(1):68-70.
15 Recommendations for Screening Any EnterobacteriaceaeAt least one carbapenem with an increased MIC (ertapenem is most sensitive, but not specific) and at least one resistant 3rd or 4th generation cephalosporins:CeftriaxoneCeftazidimeCefotaximeCeftizoximeCefepimeScreen for KPC, MBL, and AmpCESBL testing is performed by automated instrumentations.
16 KPC Klebsiella pneumoniae Carbapenemase 1st reported on the East Coast of the United States in the late 1990’sCurrently found worldwide. Can be associated with other Enterobacteriaceae.KPC gene is plasmid-mediated which has contributed to the rapid dissemination across the globe.Hydrolyze all beta-lactam antibiotics.Can be detected byModified Hodge TestBoronic Acid inhibition
17 Modified Hodge Test (MHT) Used for the detection of KPC producing isolatesCarbapenem disk: meropenem – gold standard – most specificIncreased MIC for imipenem seen is Proteus, Providencia, and MorganellaIndentation of the inhibition zone indicates that the test strain is hydrolyzing the carbapenem.Not specific for KPCAmpC enzymes can give weak false positive results that can be accentuated by porin loss
19 Alternative Test Rosco kit for carbapenemase detection 4 disks all on one plate:Meropenem (MRP10)Meropenem + Boronic Acid (MRPBO) *KPCMeropenem + Cloxacillin (MR+CX) *AmpCMeropenem + DPA Dipicolinic acid (MR+DP) *MBLEasier to read compared to the MHTReduced the risk of false positives for KPCDetects true hyperproduction of AmpC by utilizing a carbapenem instead of a cephamycin
22 Metallo Beta-Lactamase (MBL) Uses a zinc cation for the hydrolysis of the beta-lactam ringActivity is inhibited by EDTA (similar to clavulanic acid and ESBL)Hydrolyze all beta-lactam antibiotics except aztreonam.Chromosomal presence found in Stenotrophomonas, Aeromonas, and Chryseobacterium.Clinically significant MBLs have transmitted to other bacterial pathogens.
24 AmpC Chromosomal = MY SPACE bugs (Morgenella, Y. enterocolitica, Serratia, Providencia, Aeromonas, Citrobacter, Enterobacter)Inducible = Any MYSPACE or Enterobacteriaceae containing AmpC plasmidOrganism may develop resistance during prolonged therapy with 3rd generation cephalosporins.Identified in organisms exhibiting the following:Resistant to cephamycinsCefoxitinCefotetanSensitive to CefepimeHyperproduction = Any EnterobacteriaceaeCaused by a mutation in the AmpC gene leading to permanent hyperproduction or derepression.
25 The AmpC gene is now found on a plasmid We can no longer predict which bacteria are AmpC positiveAmp C is upregulated by treatment with β-lactam drugsAmount of enzyme produced is dependant on the selection of stable mutants with upregulated genes.
27 AmpC Many different AmpC enzymes C. freundii clusterCMY-2Enterobacter clusterMIR-1, ACT-1M. morganii cluster**DHA-1H. alvei clusterACC-1Aeromonas clusterCMY-1 and FOX-1Degree of carbapenemase resistance is dependant on type of AmpC in a porin deficient isolate
29 Education and Training In the past, technologists did not need to use antibiotic classification in everyday use.Education may be needed to help technologists understand the importance of classifications.Tools are available in automated instrumentation that can help technologists on the bench.
32 Tools in Instrumentation Custom comments/alerts can be printed on the patient report generated from the instrument.Activation of these tools can significantly increase the technologist’s awareness on when to appropriately screen for carbapenemases.Can also be used for many other difficulties in AST reporting.