Pseudomonas aeruginosa ; ; - -lactamases lactamases - - PowerPoint PPT Presentation

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Pseudomonas aeruginosa ; ; - -lactamases lactamases - - PowerPoint PPT Presentation

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Pseudomonas aeruginosa ; ; - -lactamases lactamases Pseudomonas aeruginosa and other enzymes enzymes and other Pr. P. Nordmann Nordmann Pr. P. hpital de Bictre, AP Bictre, AP- -HP HP hpital de Facult de M

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SLIDE 1

Pseudomonas aeruginosa Pseudomonas aeruginosa; ; ß ß-

  • lactamases

lactamases… …

and other and other enzymes enzymes

  • Pr. P.
  • Pr. P. Nordmann

Nordmann hôpital de hôpital de Bicêtre, AP Bicêtre, AP-

  • HP

HP Facult Faculté é de M de Mé édecine Paris decine Paris-

  • Sud, Universit

Sud, Université é Paris XI, Paris XI, K. K.-

  • Bicêtre, France

Bicêtre, France

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SLIDE 2
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SLIDE 3

Efflux Outer membrane PBP

  • Outermembrane

permeability (100-fold less than that of E. coli)

  • Efflux : at least twelve

systems...

  • Naturally-occurring

cephalosporinase and

  • xacillinase

Enzymes

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Cytoplasm Cytoplasm Cytoplasm

Inner (cytoplasmic) membrane Outer Membrane Lipopolysaccharide Porin Braun lipoprotein

Gram Gram-

  • negatives

negatives

Peptidoglycan

ß-Lactam resistance = +++ ß-lactamases

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SLIDE 5

Wild-type P. aeruginosa

Natural resistance Natural resistance : : ! ! aminop aminopé énicillines nicillines ; ; aminop aminopé énicilline nicilline/a. /a. clav clav. . ! !C1G, C2G C1G, C2G ! ! C3G ( C3G (cefotaxime cefotaxime, , ceftriaxone ceftriaxone) ) ! ! ertap ertapé én nè ème me Natural Natural susceptibility susceptibility: : ! ! carboxypenicillins carboxypenicillins ( (ticarcillin ticarcillin) ) ! ! ureidopenicillins ureidopenicillins ( (piperacillin piperacillin) ) ! ! cephalosporins cephalosporins ( (ceftazidime ceftazidime, , cefsulodin cefsulodin, , cefepime cefepime, , cefpirome cefpirome) ) ! ! monobactam monobactam ( (aztreonam aztreonam) ) ! ! carbapenems carbapenems ( (imipenem imipenem, , meropenem meropenem) )

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Acquired resistance Acquired resistance to to ß ß-

  • lactams

lactams; ; ß ß-

  • lactamases

lactamases

! ! Overexpression Overexpression of

  • f cephalosporinase

cephalosporinase ! ! Restricted Restricted-

  • spectrum penicillinases

spectrum penicillinases ! ! Expanded Expanded-

  • spectrum

spectrum ß ß-

  • lactamases

lactamases ( (ESBLs ESBLs) ) ! ! Carbapenemases Carbapenemases ( (metallo metallo-

  • ß

ß-

  • lactamases

lactamases) ) ! ! Expanded Expanded-

  • spectrum oxacillinases

spectrum oxacillinases

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SLIDE 7

In-vitro spectrum of ß-lactamases In-vitro spectrum of ß-lactamases

Pénicillinss Cephalosporinss 1st et 2nd generation* ß-lactamins/ Inhibitors of ß-lactamases Carbapenemss Cephalosporins 3rd génération cefepime**, cefpirome**

Enzyme ESBLs Overexpression of cephalosporinases Carbapenemases (metallo-enzymes)

Class

Penicillinases Cephalosporinases

A B C D

Oxacillinases

* Cephamycins excluded for ESBLs ** Cefepime, cefpirome excluded for overexpressed cephalosporinase

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SLIDE 8

Overexpressed cephalosporinases

! ! Resistance Resistance to all to all β β-

  • lactams except

lactams except to to carbapenems carbapenems ! !9 9-

  • 12 % of

12 % of the strains the strains

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SLIDE 9

Narrow Narrow-

  • spectrum penicillinases

spectrum penicillinases

! ! Carbenicillinases Carbenicillinases (PSE), oxacillinases (OXA), TEM (PSE), oxacillinases (OXA), TEM resistance resistance to : to : ticarcillin ticarcillin, , piperacillin piperacillin, , cefsulodin cefsulodin intermediated susceptibility intermediated susceptibility to : to : cefepime cefepime, , cefpirome cefpirome, , aztreonam aztreonam susceptibility susceptibility to : to : ceftazidime ceftazidime, imipenem, , imipenem,meropenem meropenem ! ! 10 % of 10 % of P. aeruginosa

  • P. aeruginosa strains

strains ( 90 % ( 90 % with with PSE PSE-

  • 1).

1). ! ! Non Non-

  • inhibitory activity

inhibitory activity of

  • f inhibitors against

inhibitors against oxacillinases

  • xacillinases and

and PSE PSE-

  • 1

1

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SLIDE 10

OXA-4

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SLIDE 11

Extended Extended-

  • spectrum

spectrum ß ß-

  • lactamases; ESBL

lactamases; ESBL (Ambler class A) (Ambler class A)

! ! TEM, SHV, VEB, PER, GES, BEL TEM, SHV, VEB, PER, GES, BEL ressitance ressitance to all to all ß ß-

  • lactams except

lactams except to to carbapenems carbapenems inhibition by inhibition by clavulanic acid clavulanic acid, , tazobactam tazobactam, , sulbactam sulbactam ! ! An exception : GES An exception : GES-

  • 2

2 Additional resistance Additional resistance to to carbapenems carbapenems

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SLIDE 12

PER-1 VEB-1

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SLIDE 13

BEL-1

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SLIDE 14

ESBLs in P. aeruginosa

Detection Distribution in France

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SLIDE 15
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SLIDE 16

P.

  • P. aeruginosa

aeruginosa: GES : GES-

  • 2

2

GES-1/GES-2: Gly170Asn 170 170

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SLIDE 17

ESBLs and P. aeruginosa

  • ESBL

ESBL genes associated with genes associated with insertion insertion sequence and sequence and transposons (SHV, transposons (SHV, PER PER-

  • 1) ,

1) ,with with class 1 class 1 integron and specific integron and specific sequences sequences ( (Re Re); VEB, GES, ); VEB, GES, BEL BEL-

  • 1

1

intI1 intI1 intI1 intI1

5'CS 5'CS 3'CS 3'CS 5'CS 5'CS

(P2) Pant ant

3'CS 3'CS Gene Gene-

  • cassette

cassette

  • Consequences

Consequences : : co co-

  • expression ; co

expression ; co-

  • selection

selection; ; co co-

  • resistance

resistance

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SLIDE 18
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Metallo- ß-lactamases

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SLIDE 20
  • Resistance to all ß-

lactamins except to aztreonam

  • Genetic background;

plasmids, transposons, integrons…

  • Associated resistance;

aminoglycosides ++

Metallo-ß-lactamases: carbapenemases (Ambler class B)

VIM-2

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SLIDE 21
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SLIDE 22
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SLIDE 23
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SLIDE 24

Heterogeneous Heterogeneous expression of expression of metallo metallo-

  • enzymes

enzymes

(Senda et al. Antimicrob. Agents Chemother. 1996 ; 40, 349-353)

MICs MICs ( (µ µg/ml) g/ml) IMP IMP MPM MPM CAZ CAZ AZT AZT P.

  • P. aeruginosa

aeruginosa MNA 1456 MNA 1456 >128 >128 >128 >128 >128 >128 >128 >128 MNA 14102 MNA 14102 >128 >128 >128 >128 >128 >128 64 64 MNA 14115 MNA 14115 32 32 64 64 >128 >128 16 16 MNA 1560 MNA 1560 2 2 16 16 >128 >128 32 32

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Metallo-β-Lactamases: epidemiology

Senda et al., Antimicrob. Agents Chemother. 1996; 40:349 Kurokawa et al., Lancet 1999; 354:955 Yamasaki et al., J. Antimicrob. Chemother. 2003; 51:631

Country Isolates Year(s) Prevalence Species

Japan 92-94

  • P. aeruginosa

3700 (17 hosp.) 0.4% (IMP) Japan 96-97

  • P. aeruginosa
  • S. marcescens

2533 3222 1.3% (IMP) 4.4% (IMP) Japan 98 & 00

  • P. aeruginosa

Enterobacteriaceae 200 800 0.7% (IMP) Greece 98-00 Pseudomonas spp. 233 (1 hosp.) 0.9% (VIM) Italy 01

  • P. aeruginosa

506 (1 hosp.) 0.8% (VIM)

Nationwide Regional

Tsakris et al., Clin. Microbiol. Infect. 2003; 9:846 Luzzaro et al., Diagn. Microbiol. Infect. Dis. 2004; 48:13 Jones et al., Diagn. Microbiol. Infect. Dis. 2004; 49: 289 Kimura et al., J. Clin, Microbiol. 2005; 43: 458

Japan 98-02 Gram-negatives 801 (3 hosp.) 1.1% (IMP) Japan 02

  • P. aeruginosa

594 1.9% (IMP/VIM)

Nationwide

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SLIDE 26

Metallo-β-Lactamases: Impact on resistance

Senda et al., Antimicrob. Agents Chemother. 1996; 40:349 Hirakata et al., Antimicrob. Agents Chemother. 1998; 42:2006 Shibata et al., J.Clin.Microbiol. 2003; 41:5407

Country Phenotype Year(s) Prevalence Species

Japan 92-94

  • P. aeruginosa

IMI-R CAZ-R 11% (IMP) Japan 91-96

  • P. aeruginosa

Enterobacteriaceae CAZ-R 25% (IMP) 32% (IMP) Japan 01-02

  • P. aeruginosa

Other GNNFs Enterobacteriaceae CAZ-R PER/SUL-R 62% (IMP) 11% (VIM) Korea 00-01 Pseudomonas spp. Acinetobacter spp. IMI-R/I 11% (VIM) 14% (VIM/IMP) Greece 01

  • P. aeruginosa

IMI-R/I 62% (VIM) Italy 99-02

  • P. aeruginosa

IMI-R 39% (VIM/IMP)

Lee et al., Emerg. Infect. Dis.. 2003; 9:868 Giakkoupi et al., J.Clin.Microbiol. 2003; 41:822 Toleman et al., J. Antimicrob. Chemother. 2005; 55:61

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SLIDE 27

Associated resistance….

Nordmann P, Poirel L. Emerging carbapenemases in Gram-negative aerobes. Clin Microbiol Infect. 2002 . Walsh TR, Toleman MA, Poirel L, Nordmann P. Metallo ß-lactamases; the quiet before the storm. Clin Microbol Rev 2005

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Extended-spectrum oxacillinases (Ambler class D)

ß-Lactamase

TIC TCC PIP TAZ CAZ FEP ATM IMP OXA-2 128 64 32 4 16 2 2 4 OXA-15 128 64 32 8 128 4 8 2 OXA-32 256 64 32 16 128 8 32 4 OXA-10 512 64 2 4 16 2 OXA-11 512 64 512 4 128 2 OXA-14 512 256 64 32 512 64 16 2 OXA-16 128 64 32 16 128 32 4 1 OXA-13 256 256 32 32 2 8 1 OXA-19 256 256 64 64 256 16 1 OXA-28 128 128 16 16 256 16 32 0,25 OXA-18 256 64 64 32 128 16 256 2 OXA-31 512 512 256 256 2 256 32 TIC, ticarcillin, TCC, ticarcillin + clav.; PIP, piperacillin; TAZ, piperacillin + tazobactam; CAZ, ceftazidime; FEP, cefepime; ATM, aztreonam, IMP, imipenem

Detection ? Epidemiology ?

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SLIDE 29

Diversity of oxacillinase resistance patterns

OXA-32 OXA-31

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Aminoglycoside resistance

  • Efflux
  • Outer membrane permeability defect
  • Modification enzymes
  • Target alteration
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APH(3')

phosphorylation

  • P. aeruginosa; natural resistance APH 3’

AAC(3)

acetylation

4'

ANT(2")

nucleotidylation

HO

3 6'

HO H2N O OH HO

2"

O HO OH H2N H2N O O NH2 NH2

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SLIDE 32
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SLIDE 33
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SLIDE 34

Novel mechanism of resistance to rifampicin

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Conclusion Conclusion

! ! Emerging mechanisms Emerging mechanisms of

  • f resistance

resistance; ESBL, ; ESBL, metallo metallo-

  • ß

ß-

  • lactamases,

lactamases, novel novel aminoglycoside resistance genes aminoglycoside resistance genes… … ! ! Novel genetic elements Novel genetic elements for for their mobility their mobility ! ! Environmental reservoir Environmental reservoir of

  • f resistance genes

resistance genes ? ? ! ! Increased Increased rate of rate of multidrug multidrug-

  • resistant

resistant P. aeruginosa

  • P. aeruginosa worldwide

worldwide

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SLIDE 36

Thanks

  • D. Aubert, PhD
  • D. Girlich, tech.
  • T. Naas, MCU-PH
  • L. Poirel, CR1 Inserm