The Beta-Lactamase Family: Classification, Detection, and

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The Beta-Lactamase Family: Classification, Detection, and Powered By Docstoc
					       The Beta-Lactamase Family:
Classification, Detection, and Interpretive
                  Criteria
        COL Helen Viscount, PhD, D(ABMM)
        LTC Steven Mahlen, PhD, D(ABMM)
Transplant patient
 Extremely resistant Klebsiella         Ampicillin: R
   pneumoniae recovered                  Pip/tazo: R
    Sensitive only to colistin and
       gentamicin
                                         Ceftazidime: R
 Patient put in isolation
                                         Ceftriaxone: R
 Isolate transmitted to 10 other
                                         Cefepime: R
  patients                               Imipenem: R
 Outcomes:                              Meropenem: R
      4/5 with bacteremia died          Aztreonam: R
      1 other died                      Amikacin: R
      2 with renal failure              Tobramycin: R
      Only 4/11 discharged without      Trimeth/sulfa: R
       renal failure
                                         Fluoroquinolones: R
                                         Gentamicin: S
                                         Colistin: S
Nursing home resident
 83 years old                       Ampicillin: R
 Pneumonia                          Pip/tazo: S
    Admitted to ICU                 Cefazolin: R
    Started on ceftriaxone and      Ceftazidime: I
     levofloxacin
                                     Ceftriaxone: S
 Blood cultures +
    K. pneumoniae                   Cefepime: S
 Based on sensi’s:                  Imipenem: S
    No more levo                    Aztreonam: S
    Kept on ceftriaxone             Tobramycin: S
 Patient got worse                  Trimeth/sulfa: R
    Had to be ventilated            Levofloxacin: I
                                     Ciprofloxacin: I
                                     Gentamicin: S
Objectives
 At the end of this workshop
  the attendee should be able to
  distinguish ESBL positive
  from carbapenemase-
  producing bacteria
 At the end of this workshop
  the attendee should be able to
  describe a method to screen
  for ESBLs
 At the end of this workshop
  the attendee should be able to
  interpret the results of the
  modified Hodge Test
Beta-lactam antibiotics
 Penicillins
   Ampicillin
   Amoxicillin
   Piperacillin
 Cephalosporins (generations)
   1st gen: cephalothin
   2nd gen (cephamycins): cefoxitin, cefotetan
   3rd gen: ceftazidime, cefotaxime, ceftriaxone
   4th gen: cefepime
Beta-lactam antibiotics
 Monobactam: aztreonam
 Carbapenems:
   Imipenem
   Meropenem
   Ertapenem
 Inhibitors
   Sulbactam (ampicillin/sulbactam: Unasyn)
   Tazobactam (piperacillin/tazobactam: Zosyn)
   Clavulanate (amoxicillin/clavulanate: Augmentin)
Mechanisms of Resistance
 Altered target (Gram negative/positive)
 Altered permeability (Gram negative)
 Production of inactivating enzymes (Gram negative/positive)
                   Gram-negative cell          Gram-positive cell




Outer membrane



                                                               Peptidoglycan
Peptidoglycan
Penicillin
Binding proteins
(PBPs)



                        Inner (cytoplasmic) membrane
Alteration of Target
 Resistance to -lactams via altered penicillin-binding
  proteins (PBPs)
   MRSA
 Vancomycin resistance in enterococci
 Fluoroquinolone resistance
Altered Permeability
 Passive diffusion of Gram-negative cell wall
 Mutate outer membrane proteins
 Active efflux
Active Efflux
Production of Inactivating Enzymes
 Chloramphenicol acetyltransferase
 Aminoglycoside-modifying enzymes
 -Lactamases
-Lactamases
 Well over 340 different enzymes
 Extended spectrum -lactamases (ESBLs)
 AmpC -lactamases
   Chromosomal
   Plasmid-mediated
 Carbapenemases
-Lactamases
 First -lactamase identified: AmpC beta-lactamase
   1940, Escherichia coli
   1940, penicillinase, Staphylococcus aureus
 First plasmid-mediated -lactamase: TEM-1
   1965, Escherichia coli, Greece
-Lactamase Activity
                       H   H
                                   -lactam
                               S
          R-CONH       C   C             CH3



                       C   N          CH3
                   O

                               COOH
Enzyme-Ser-OH
-Lactamase Activity
                       H   H

                               S
     R-CONH            C   C           CH3



              O        C   N          CH3



                       O   H   COOH
   HOH
                  Ser

              Enzyme
L
                                                                
    L L                             
                                                   
                                       
  L                    
                                            
             L        L                                 

                                             
                     L
                                                               
-lactamase
                                 L                    
production          L                          
Types of Beta-Lactamases
 ESBLs
 AmpCs
 Carbapenemases
ESBLs
ESBLs
 Extended-spectrum beta-lactamases (ESBLs) are mutant
  enzymes with a broader range of activity than their parent
  molecules
 They:
   Hydrolyze 3rd and 4th gen cephalosporins and aztreonam
   Do not affect cephamycins (2nd gen ceph) or carbapenems
   Remain susceptible to beta-lactamase inhibitors
ESBLs
 The most common plasmid-mediated ß-lactamases in
  Enterobacteriaceae are TEM-1, TEM-2, and SHV-1
   TEM: Escherichia coli
     Named after first patient with a urinary tract infection that was not
      treatable with ampicillin
     Her name: Temorina
   SHV: Klebsiella pneumoniae
     “Sulfhydryl variant”; amino acids in the enzyme that cross-link with other
      molecules
 “Classical” ESBLs are derived from TEM and SHV enzymes
 “Non-classical” ESBLs are derived from enzymes other than
  TEM or SHV
Classical ESBLs
 Primarily found in E. coli and Klebsiella spp.
 Differ from their parent TEM or SHV enzymes by only 1-4
  amino acids
 >100 TEM- or SHV-derived beta-lactamases have been
  described – most are ESBLs
Non-classical ESBLs
 Many described, but less common than classical ESBLs
   CTX-M
     Found in multiple genera of Enterobacteriaceae
     Preferentially hydrolyze cefotaxime
     U.S., Europe, South America, Japan, Canada
   OXA
     Mainly in P. aeruginosa
     Primarily hydrolyze ceftazidime
     France, Turkey
ESBL Epidemiology
 ESBLs first appeared in Europe in the mid-1980s
 Worldwide, but prevalence varies widely geographically and
  between institutions
 U.S. national average for ESBLs in Enterobacteriaceae ~3%
ESBL Epidemiology
 ESBL producers especially prevalent in ICUs and long
  term care facilities
   Becoming more widespread in the community also
 Have been associated with outbreaks
   Typically arise in ICU
     Plasmid transfer between GNRs
     Organism transfer between patients
   Control of outbreaks
     Infection control practice – isolation
     Restriction of 3rd and 4th generation cephalosporins
     Antimicrobial cycling
Clinical Significance

 Despite appearing susceptible to one or more penicillins,
  cephalosporins, or aztreonam in vitro, the use of these agents
  to treat infections due to ESBL-producers has been associated
  with poor clinical outcome
Clinical Significance
 ESBL genes are often carried on plasmids that also
  encode resistance to multiple classes of antimicrobials
   Aminoglycosides, Fluoroquinolones
   Trimethoprim/Sulfamethoxazole
 Treatment experience is largely based on classical ESBL
  producers
   Carbapenems
   ß-lactam/inhibitor combinations
Typical ESBL Susceptibility Profile
 Amp: R                   Amp: R
 Piperacillin: R          Piperacillin: R
 Pip/tazo: S              Pip/tazo: S
 Cefazolin: R             Cefazolin: R
 Cefoxitin: S             Cefoxitin: S
 Ceftazidime: S           Ceftazidime: R
 Ceftriaxone: R           Ceftriaxone: R
 Cefepime: R              Cefepime: R
 Aztreonam: S             Aztreonam: R
 Imipenem/meropenem: S    Imipenem/meropenem: S
AmpCs
AmpC: General
 Chromosomal
   Escherichia coli
   Citrobacter freundii
   Enterobacter aerogenes, E. cloacae
   Serratia marcescens
   Morganella morganii
   Hafnia alvei
   Providencia rettgeri, P. stuartii
   Pseudomonas aeruginosa
   Aeromonas sp.
AmpC: General
 Are not inhibited by -lactamase inhibitors
 Normally are repressed, so produced at low levels
 Chromosomal: inducible
   In all except E. coli
   In the presence of certain -lactam antibiotics
   Normally, produced at low levels
 Plasmid-mediated also
The AmpC of E. coli
 Chromosomal, but not          Amp: S
  inducible                     Amox/clav: S
 Normally expressed at low     Piperacillin: S
  levels                        Pip/tazo: S
 Regulated by a growth         Cefoxitin: S
  rate-dependent attenuation    Ceftazidime: S
  mechanism                     Ceftriaxone: S
 Can become highly             Cefepime: S
  expressed with mutations      Aztreonam: S
                                Imipenem/meropenem: S
AmpC Induction and Derepression
 Is induction clinically relevant?
 True danger—mutation in induction pathway
   “Derepressed mutant”
   150-1000 fold more enzyme produced than normal
Chromosomal AmpC profile
 Normal                       Derepressed profile
     Amp: R                       Amp: R
     Amox/clav: R                 Amox/clav: R
     Piperacillin: S              Piperacillin: R
     Pip/tazo: S                  Pip/tazo: R
     Cefoxitin: R                 Cefoxitin: R
     Ceftazidime: S               Ceftazidime: R
     Ceftriaxone: S               Ceftriaxone: R
     Cefepime: S                  Cefepime: S
     Aztreonam: S                 Aztreonam: R
     Imipenem/meropenem: S        Imipenem/meropenem: S
Plasmid-Mediated AmpCs (pAmpC)
 First true proof of AmpC on plasmid: 1988
    MIR-1, found in Klebsiella pneumoniae
    90% identical to E. cloacae ampC
 Some are also inducible (DHA-1)
 Most frequently found in K. pneumoniae
 Also commonly found in:
    K. oxytoca
    Salmonella sp.
    P. mirabilis
 E. coli, E. aerogenes also
pAmpCs: Distribution
 World-wide distribution
   Africa, Asia, Europe, Middle East, North America, South
    America, Central America
 CMY-2 is most prevalent globally
   Algeria, France, Germany, Greece, India, Pakistan, Taiwan,
    Turkey, UK, US
  ESBLs vs AmpCs

                            ESBLs   AmpCs

Inhibitors (pip/tazo,
amp/sulbactam, amox/clav)     S       R

Cefoxitin, cefotetan          S       R

Ceftazidime,
                             R        R
ceftriaxone

Cefepime                    S/R       S
Carbapenemases
Carbapenemases
 Carbapenem resistance:
   High level production of chromosomal AmpC with decreased
    outer membrane permeability (porins)
     E. cloacae, E. aerogenes
     C. freundii
     E. coli
     S. marcescens
     K. pneumoniae (porins)
Carbapenemases
 Carbapenem resistance:
   Changes in affinity of PBPs for carbapenems
   Carbapenemases
 Frequently, bugs that produce a carbapenemase produce
  other -lactamases
Carbapenemases
 KPC (plasmid, K. pneumoniae)
    “Klebsiella pneumoniae carbapenemase”
 IMI-1 (plasmid, E. cloacae)
 Nmc-A (plasmid, E. cloacae)
 Sme-1 (plasmid S. marcescens)
 IMP-1 (plasmid, S. marcescens, P. aeruginosa)
 L-1 (chromosomal, Stenotrophomonas maltophilia)
Carbapenemases: Profile
 R to carbapenems, penicillins, cephalosporins
 S or R to aztreonam, depending on enzyme
 So the key:
   Look for intermediate or R to imipenem or meropenem!
KPC
 Infection control emergency!!!
   May test sensitive to carbapenems though!
 Extensive multidrug resistance (XDR)
 Very rapid spread
 Empiric therapy: colistin + tigecycline
 KPC 1-8
Further reading
 Yang, 2007. Ann. Pharmocother. 41:1427-1435
 Jacoby, 2009. Clin. Microbiol. Rev. 22:161-182
 Black et al, 2005. J. Clin. Microbiol. 43:3110-3113
 Livermore et al, 2001. J. Antimicrob. Chemother. 48 Suppl
  1: 87-102
 Pfaller and Segreti, 2006. Clin. Infect. Dis. 42: S153-163.

				
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