Resistance to b-lactam antibiotics within the Enterobacteriaceae - PowerPoint PPT Presentation

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Resistance to b-lactam antibiotics within the Enterobacteriaceae

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Title: Resistance to b-lactam antibiotics within the Enterobacteriaceae


1
  • Resistance to b-lactam antibiotics within the
    Enterobacteriaceae
  • Paul D. Fey, Ph. D.
  • University of Nebraska Medical Center

2
Objectives
  • Understand the mechanisms of ?-lactam resistance
    in GNRs
  • Understand how ?-lactamases are transferred.
  • Know the characteristics of three classes of
    ?-lactamases
  • ESBLs
  • AmpC
  • KPC

3
Mechanism of ?-Lactam Action
  • Bactericidal
  • ?-lactams bind and inhibit penicillin binding
    proteins (PBPs)
  • PBPs are responsible for assembly, maintenance,
    and regulation of peptidoglycan (cell wall)
    metabolism.
  • Disruption of peptidoglycan synthesis

4
Mechanisms of GNR Resistance to ?-lactams
  • Porin-mediated resistance
  • Antibiotic does not reach target.
  • b-lactamase
  • Majority of resistance to b-lactam antibiotics
    mediated through b-lactamases.
  • Many different types of b-lactamases with
    different substrate (antibiotic) specificities.

5
Common ?-Lactamases in GNRs
NEJM 3524. 2005
6
How are b-lactamases transferred?
  • Transfer of Plasmids.
  • Extrachromosomal DNA
  • Usually carry antibiotic resistance genes
  • These genes can be encoded on transposons, which
    are also mobile.
  • TEM-1 has been transferred between the
    Enterobacteriaceae and H. influenzae and the
    Neisseriaceae

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12
b-lactam antibiotics
  • Penicillins
  • Ampicillin
  • Piperacillin
  • Beta-lactam/beta-lactamase inhibitors
  • Ampicillin/sulbactam
  • Amoxicillin/clavulanate
  • Ticarcillin/clavulanate
  • Piperacillin/Tazobactam

13
b-lactam antibiotics
  • First Generation cephalosporins
  • Cefazolin
  • Cephalothin
  • Second Generation oral antibiotics
  • Cefuroxime (many others)
  • Second Generation cephamycins
  • Cefoxitin
  • Cefotetan

14
b-lactam antibiotics
  • Third generation cephalosporins
  • Cefotaxime
  • Ceftriaxone
  • Ceftazidime
  • Fourth generation cephalosporins
  • Cefepime
  • Monobactams
  • Aztreonam

15
b-lactam antibiotics
  • Carbapenems
  • Imipenem
  • Meropenem
  • Ertapenem
  • Doripenem

16
ESBLs
  • Enzymes capable of hydrolyzing third-generation
    cephalosporins.
  • Plasmid-mediated
  • Derivatives (mutants) of original TEM-1 and SHV-1
    b-lactamases.
  • Susceptible in-vitro to clavulanate and
    cefoxitin.

17
E. coli susceptibility Report
  • Ampicillin R
  • Piperacillin R
  • Cephalothin R
  • Cefoxitin S
  • Cefotaxime R
  • Ceftazidime I
  • Ceftriaxone R
  • Aztreonam I
  • Cefepime S
  • Pip/Tazo I
  • Imipenem S

18
Laboratory detection of ESBLs
  • Resistance or intermediate to third-generation
    cephalosporins.
  • Cefoxitin and cefotetan susceptible.
  • ESBL disk diffusion test (clavulanate inhibition)
  • E-test ESBL strip
  • Confirmatory ESBL MIC test (Microscan)
  • K. pneumoniae, K. oxytoca, E. coli, P. mirabilis

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E. coli ESBL susceptibility report
  • Ampicillin R
  • Piperacillin R
  • Cephalothin R
  • Cefoxitin S
  • Cefotaxime R
  • Ceftazidime I?R
  • Ceftriaxone R
  • Aztreonam I?R
  • Cefepime S?R
  • Pip/Tazo I
  • Imipenem S

21
Enterobacter cloacae susceptibility report
  • Ampicillin R
  • Piperacillin R
  • Cephalothin R
  • Cefoxitin R
  • Cefotaxime R
  • Ceftazidime I
  • Ceftriaxone R
  • Aztreonam I
  • Cefepime S
  • Pip/Tazo R
  • Imipenem S

22
AmpC b-lactamases
  • Chromosomally encoded-cell wall turnover
  • Enterobacter sp., Citrobacter sp., Serratia sp.,
    Morganella sp. Even E. coli.
  • Third-generation cephalosporins are not good
    inducers of AmpC b-lactamase
  • Third-generation cephalosporin resistant strains
    are derepressedmeaning that the AmpC b-lactamase
    is not inducible anymore.
  • AmpC mutants are cephamycin resistant

23
E. coli susceptibility report
  • Ampicillin R
  • Piperacillin R
  • Cephalothin R
  • Cefoxitin R
  • Cefotaxime R
  • Ceftazidime I
  • Ceftriaxone R
  • Aztreonam I
  • Cefepime S
  • Pip/Tazo R
  • Imipenem S

24
  • What do you do?

25
Other concepts to know about AmpC b-lactamases
  • They are transferred on plasmids as well.
  • CMY, LAT, BIL, MOX, ACC, FOX, DHA
  • Almost all ceftriaxone-resistant Salmonella
    isolated in the United States carry a
    plasmid-mediated AmpC b-lactamase called CMY-2.
  • E. coli UTI isolates carry plasmid-mediated AmpC
    b-lactamases

26
Mechanisms of Carbapenem Resistance
  • Carbapenemase hydrolyzing enzymes
  • Porin loss OprD
  • ESBL or AmpC porin loss

27
Carbapenemases
  • The most versatile family of ?-lactamases
  • Two major groups based on the hydrolytic
    mechanism at the active site
  • Serine at the active site class A and D
  • Zinc at the active site class B
  • All carbapenemases hydrolyze penicillins,
    extended spectrum cephalosporins, and carbapenems

28
Carbapenemase Classification
Molecular Class A B D
Functional Group 2f 3 2d
Aztreonam Hydrolysis - -
EDTA Inhibition - -
Clavulanate Inhibition - ?
29
Klebsiella pneumoniae
  • Ampicillin R
  • Piperacillin R
  • Cephalothin R
  • Cefoxitin S
  • Cefotaxime R
  • Ceftazidime I
  • Ceftriaxone R
  • Aztreonam I
  • Cefepime S
  • Pip/Tazo R
  • Imipenem I
  • Might need to screen for carbapenemase

30
Carbapenemases Class A
  • First identified 1982 in UK
  • Four major families
  • Chromosomally encoded
  • Serratia marcescens enzyme (SME)
  • Not metalloenzyme carbapenemases (NMC)
  • Imipenem-hydrolyzing ?-lactamases (IMI)
  • Plasmid encoded
  • Klebsiella pneumoniae carabapenemases (KPC)
  • Guiana Extended-Spectrum (GES)

31
KPC
  • Molecular class A and functional group 2f
  • Inhibited by clavulanic acid but not by EDTA
  • Confers resistance to ALL ?-LACTAM antibiotics
  • Plasmid-encoded
  • Associated with other resistant genes
    (aminoglycosides, fluoroquinolones)
  • Transferable

32
KPC Epidemiology
  • Predominantly in K. pneumoniae (KP)
  • Reported in Enterobacter spp., Salmonella spp.,
    E. coli, P. aeruginosa, and Citrobacter spp.
  • First identified in KP clinical isolate from
    North Carolina in 1996 (KPC-1)
  • KPC-2, -3, and -4 have been reported.
  • Mostly identified on the East cost

33
KPC Epidemiology
  • KPC producers have been identified outside USA
  • France
  • Brazil
  • Columbia
  • China
  • Not detected at the University of Nebraska
    Medical Center
  • 45 ESBL-like isolates collected-6 had elevated
    carbapenem MICs-none contained KPC

34
When to Suspect a KPC Producer
  • Enterobacteriaceae
  • Resistance to extended spectrum cephalosporins
    (cefotaxime, ceftazidime, and ceftriaxone)
  • Variable susceptibility to cephamycins
    (cefoxitin, cefotetan)
  • Carbapenem MICs ? 2 ?g/ml

35
How to Detect a KPC Producer
  • Antimicrobial susceptibility tests (ASTs)
  • MIC
  • Carbapenem MIC ? 2 ?g/ml
  • Disk diffusion
  • Carbapenem I or R
  • Among carbapenems, ertapenem
  • Most sensitive
  • less specific

Anderson et al. 2007. JCM 45 (8) 2723
36
How to Detect a KPC Producer
  • Commercial systems
  • Inconsistent detection of KPC-producing isolates
  • Tenover et al. 2006. EID. 121209-1213
  • Breakpoints do not match CLSI recommendations

37
Definitive ID of a KPC Producer
  • Modified Hodge test
  • 100 sensitivity to detect KPC
  1. Swab E. coli ATCC 25922 onto plate to create lawn
    Place imipenem disk in center.
  2. Streak test isolates from edge of disk to end of
    plate.
  3. Incubate overnight.
  4. Look for growth of E. coli around test isolate
    streak - indicates carbapenem-hydrolyzing enzyme.

pos
pos
pos
neg
neg
neg
meropenem
ertapenem
imipenem
Janet Hindler, Whats New in the 2008 CLSI
Standards for (AST)?
38
Definitive ID of a KPC Producer
  • PCR
  • The method of choice to confirm KPC

39
Alternative Treatment for a KPC Producer
  • Tigecycline (100.0 effective)
  • Colistin (88.1 effective)
  • SENTRY report. AAC. 2008. Feb52(2)570-3
  • Minocycline
  • A strategy for susceptibility testing is needed

40
Conclusions
  • ESBL detectionCLSI guidelines present
  • Need to have guidelines to detect ESBLs present
    in other species besides E. coli, K. pneumoniae,
    K. oxytoca, and P. mirabilis.
  • AmpC detection-No guidelines available
  • KPC detection-Not widespread, need to have lower
    concentrations of carbapenems on panels.
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