Cephalosporin Resistant Escherichia coli

Dr.T.V.Rao MD

Dr.T.V.Rao MD 1

Beta-lactam antibioticsPenicillins

Ampicillin

Amoxicillin

Piperacillin

Cephalosporins (generations)

1st gen: cephalothin

2nd gen (Cephamycins): cefoxitin, cefotetan

3rd gen: ceftazidime, cefotaxime, ceftriaxone

4th gen: cefepimeDr.T.V.Rao MD 2

Dr.T.V.Rao MD3

Definition of beta lactamases

Beta lactamases are enzymes produced by some gram-positive and gram-negative bacteria that hydrolyze beta lactam antibiotics

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BETA LACTAM RING

PENICILLIN

BETA LACTAM RING

CEPHALOSPORIN

BETA LACTAMASES enzymes that inactivate the beta-lactam ring

Dr.T.V.Rao MD

The β-lactam family of antibiotics

Ceftriaxone 3rdTicarcillin

Ceftazidime 3rdMezlocillin

Cefotaxime 3rdCarbenicillin

ErtapenemCefmetazoleCefuroxime 2ndAmpicillin

Meropenem

CefotetanCefamandole 2ndMethicillin

AztreonamImipenemCefoxitinCephalothin 1stBenzyl-penicillin

MonobactamsCarbapenemsCephamycinsCephalosporinsPenicillin's

Cefepime 4th

Dr.T.V.Rao MD 5

Other Beta-lactam antibiotics

Monobactam: aztreonam

Carbapenems:

Imipenem

Meropenem

Ertapenem

Inhibitors

Sulbactam (ampicillin/sulbactam: Unasyn)

Tazobactam (piperacillin/tazobactam: Zosyn)

Clavulanate (amoxicillin/clavulanate: Augmentin)Dr.T.V.Rao MD 6

What are 3rd Generation Cephalosporins

Third-generation cephalosporins are broad-spectrum drugs with high intrinsic activity against gram-negative species. The rising resistance to these drugs is worrisome because it could be a proxy for the emergence and spread of Enterobacteriaceae strains producing extended-spectrum β-lactamase

Dr.T.V.Rao MD 7

-Lactamase Activity

C C

C N

H H

R-CONH

S

COOH

CH3

CH3

O

Enzyme-Ser-OH

-lactam

Dr.T.V.Rao MD 8

-Lactamase Activity

C C

C N

H H

R-CONH

S

COOH

CH3

CH3

O

HO

Ser

Enzyme

HOH

Dr.T.V.Rao MD 9

L

L

L

LL

L

L

L Lb-lactamaseproduction

Dr.T.V.Rao MD 10

MODE OF ACTION OF BETA LACTAMS IN GRAM NEGATIVES

SUSCEPTIBLE RESISTANT

-Lactam Antibiotic

Diffusion through Porin Blocks Entry

Outer Membrane Efflux Pump

Diffusion through Beta-Lactamase

Peptidoglycan Hydolyzes Beta-Lactam

Penicillin Binding Proteins Changes in PBP results in Failure to Bind to -Lactam Cell Death

Dr.T.V.Rao MD 11

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Some beta-lactamases only inactivate a small number of antibiotics e.g.

penicillin

Others have extended spectrum to all the penicillin's and cephalosporins e.g. cefuroxime, ceftriaxone (ESBLs)

In addition may also carry resistance to other antibiotics e.g. ciprofloxacin.Dr.T.V.Rao MD

What it means as 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 inhibitorsDr.T.V.Rao MD 13

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

Dr.T.V.Rao MD 14

Non-classical ESBLs

Many described, but less common than classical ESBLs

CTX-MFound in multiple genera of EnterobacteriaceaePreferentially hydrolyze cefotaximeU.S., Europe, South America, Japan, Canada

OXAMainly in P. aeruginosaPrimarily hydrolyze ceftazidimeFrance, Turkey

Dr.T.V.Rao MD 15

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ESBLs

Extended-spectrum β-lactamases

www.lahey.org/studies/webt.htm

>180 enzymes described (119 TEM, 45 SHV)

All mutations of older TEM and SHV plasmid-mediated β -lactamases

TEM-3, TEM-4, etc.

SHV-2, SHV-3, etc.

CTX-M-1,2, etc. and Toho-type

OXA-type

PER-1 and 2Resistance conferred to extended-spectrum penicillins, 3rd

and 4th generation cephalosporins and aztreonam (not imipenem or cephamycins)

Dr.T.V.Rao MD

Extended-Spectrum β-Lactamases

β-lactamases capable of conferring bacterial resistance to

the penicillin's

first-, second-, and third-generation cephalosporins

aztreonam

(but not the Cephamycins or Carbapenems)

These enzymes are derived from group 2b β-lactamases (TEM-1, TEM-2, and SHV-1)

differ from their progenitors by as few as one AA

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Plasmid-Mediated AmpCs

B-lactamases derived from chromosomally encoded clavulanate-resistant AmpC cephalosporinases of Citrobacter, Enterobacter & Morganella spp.

Genes are typically encoded on large plasmids and carry additional resistance genes

Dr.T.V.Rao MD

Some premises

• Growing resistance to 3-gen cephalosporins

• Mostly ESBLs in E. coli & Klebsiella; AmpC in Enterobacter, Citrobacter, Serratia… but not always

• Identification of mechanism aids

– Epidemiological investigation / control– Treatment choice– Recognition of the exceptional e.g. MBLs

Dr.T.V.Rao MD 19

Resistance in E.coli a global concern

IN 2009, E coli resistance levels to third-generation cephalosporins in the United States compared favourably with those of the rest of the developed world with rates about equal to those in Scandinavian countries (Iceland, Estonia, Norway), and a lower reported resistance than the Netherland

Dr.T.V.Rao MD 20

ESBLs characterization

TEM ESBLs

- More than 30 described

- TEM-6, TEM-10, TEM-12, and TEM-26

SHV ESBLs

- More than 10

CTX-M ESBLs

AmpC

Derived from chromosomal AmpC genes of gram- negative organisms, such as Citrobacter freundii, Enterobacter cloacae, and Aeromonas spp.

Dr.T.V.Rao MD 21

Detecting ESBL producers

steps:• Screen for resistance with an indicator

cephalosporin

• Do confirmatory test on those found resistant

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Choice of indicator cephalosporin

Sensitivity Specificity

Cefotaxime & ceftazidime

Good Good

Cefpodoxime Good Moderate

Cefuroxime Poor Poor

Cephalexin or cephradine

Moderate Poor

Cefpirome or Cefepime

Poor Good

Dr.T.V.Rao MD 23

Detection of ESBLs: step 2

Seek ceph/clav synergy in ceph R isolates

Double discCombination discEtest

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Combination discs

Disc with cephalosporin

alone

Disc with cephalosporin + clavulanic

acid

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Dr.T.V.Rao MD 26

Etest for ESBLs

Cefotaxime

Cefotaxime+

clavulanateDr.T.V.Rao MD 27

ESBL – leading cause of Treatment of Failure

ESBLs are bacterial enzymes that confer resistance to many highly effective antibiotic classes that can go undetected if conventional testing methods are used in the lab, ultimately leading to treatment failure.

Dr.T.V.Rao MD 28

Resistance in Gram-negative bacteria: Enterobacteriaceae.

The emergence and spread of resistance in Enterobacteriaceae are complicating the treatment of serious nosocomial infections and threatening to create species resistant to all currently available agents. Approximately 20% of Klebsiella pneumoniae infections and 31% of Enterobacter spp infections in intensive care units in the United States now involve strains not susceptible to third-generation cephalosporins

Dr.T.V.Rao MD 29

The AmpC of E. coliChromosomal, but not

inducible

Normally expressed at low levels

Regulated by a growth rate-dependent attenuation mechanism

Can become highly expressed with mutations

Amp: S

Amox/clav: S

Piperacillin: S

Pip/tazo: S

Cefoxitin: S

Ceftazidime: S

Ceftriaxone: S

Cefepime: S

Aztreonam: S

Imipenem/meropenem: SDr.T.V.Rao MD 30

ESBLs vs AmpCs

ESBLs AmpCs

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

Cefoxitin, cefotetan S R

Ceftazidime, ceftriaxone R R

Cefepime S/R SDr.T.V.Rao MD 31

CTX-M-type ESBLs Until 2000, most ESBL producers were hospital

Klebsiella spp. with TEM and SHV mutant β-lactamases

Now, the dominant ESBLs across most of Europe and Asia are CTX-M enzymes, which originated as genetic escapes from Kluyvera spp

Currently recognized as the most widespread and threatening mechanism of antibiotic resistance, both in clinical and community settings

80% of ESBL-positive E. coli from bacteraemias in the UK and Ireland are resistant to fluoroquinolones 40% are resistant to gentamicin

Livermore, DM J. Antimicrob. Chemother 2009 Dr.T.V.Rao MD 32

Clinical Significance ESBL genes are often carried on plasmids that

also encode resistance to multiple classes of antimicrobials

Aminoglycosides, Fluoroquinolones

Trimethoprim /Sulphmethoxazole

Treatment experience is largely based on classical ESBL producers

Carbapenemsß-lactam/inhibitor combinations

Dr.T.V.Rao MD 33

ESBL – K pneumonia Resistance in K pneumoniae to third-

generation cephalosporins is typically caused by the acquisition of plasmids containing genes that encode for extended-spectrum beta-lactamases (ESBLs), and these plasmids often carry other resistance genes as well. ESBL-producing K pneumoniae and Escherichia coli are now relatively common in healthcare settings and often exhibit multidrug resistance

Dr.T.V.Rao MD 34

ESBLs: evolution in detection and reporting

Until 2009: Search for ESBL production by

specific phenotypic testing

- All confirmed ESBL-producers to be reported as RESISTANT to all PENICILLINS, CEPHALOSPORINS and AZTREONAM regardless of MICs

Dr.T.V.Rao MD 35

Enterobacteriaceae: Breakpoints revised

AgentCLSI 2009 CLSI 2010

S I R S I R

Cefazolin ≤8 16 ≥32 ≤1 2 ≥4

Cefotaxime ≤8 16-32 ≥64 ≤1 2 ≥4

Ceftriaxone ≤8 16-32 ≥64 ≤1 2 ≥4

Ceftazidime ≤8 16 ≥32 ≤4 8 ≥16

Aztreonam ≤8 16 ≥32 ≤4 8 ≥16

Cefipime ≤8 16 ≥32 ≤8 16 ≥32

Dr.T.V.Rao MD 36

ESBLs: Evolution in detection and reportingMIC (mg/L)

Antibiotic CLSI 2012

CLSI 2012 EUCAST 2012 EUCAST 2012

Cefotaxime Ceftriaxone

S≤ 1

R> 2

S≤1

R> 2

Ceftazidime 4 8 1 4

Cefepime 8 16 1 4

Aztreonam 4 8 1 4Dr.T.V.Rao MD 37

Escherichia coli ESBL+ (CTX-M-1), CLSI 2012MIC (mg/L)

Dr.T.V.Rao MD 38

Ampicillin >128 RAmoxi/Clav 32 RPip/Tazo 8 SCephalotin 32 RCefotaxime 32 RCeftazidime 1 SCefepime 8 SErtapenem 0.12 SMeropenem 0.12 SESBL positive

Amikacin 2 SGentamicin 16 RCiprofloxacin >32 RLevofloxacin >32 R

Bacteria not to test for ESBLs

Acinetobacters–Often S to clavulanate alone

S. maltophilia–+ve result by inhibition of L-2

chromosomal b-lactamase, ubiquitous in the species

Dr.T.V.Rao MD 39

Other beta-lactamases capable of hydrolyzing expanded-spectrum cephalosporins:

Some OXA-type variants (e. g. OXA-14/17)

• AmpC beta-lactamases (e. g. CMY, DHA)

• Serine carbapenemases (e. g. KPC)

• Metallo-beta-lactamases (e. g. IMP, VIM, NDM

Dr.T.V.Rao MD 40

Who are At Risk with ESBL+E.coli

Travellers 5.2 times more likely to be colonised by ESBL+ E. coli

Highest rates of ESBL carriage associated with travels to Africa and Indian subcontinent

All ESBL+ E. coli had CTX-M enzymes (71% CTX-M-15, 26% CTX-M-14)

Dr.T.V.Rao MD 41

Larger Inoculum Effect

More vulnerable to hydrolysis of β-lactamase

ESBL-KP + 3rd or 4th generation cephalosporin

How about Cephamycins? Little information

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Role of the Microbiology Lab

“ Each laboratory should have a staff member with the time, interest, and expertise to provide leadership in antibiotic testing and resistance. This person would read relevant publications, network with other laboratories, and evaluate potentially useful tests to detect new forms of resistance before new CLSI-recommended tests become available”

- Ken Thomson, Emerging Infect. Dis., 2001Dr.T.V.Rao MD

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 ICUPlasmid transfer between GNRs

Organism transfer between patients

Control of outbreaksInfection control practice – isolation

Restriction of 3rd and 4th generation cephalosporins

Antimicrobial cyclingDr.T.V.Rao MD 44

ESBL are Emerging Challenges in Patient Care for Clinicians and Microbiologists

ESBLs: complex evolution since the 1980s (multiple enzymes, High-Risk clones)

Now a very major resistance issue in enterics: globally disseminated, ubiquitous (hospital, LTCFs, community), high rates

Challenge of intestinal carriage (intra/inter institutional dissemination, cross-border transmission) and of extra-human reservoirs

Detection and reporting issues

Treatment issuesDr.T.V.Rao MD 45

Programme Created by Dr.T.V.Rao MD for Microbiologists and Health Care

Workers email

doctortvrao@gmail.com

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