This document discusses drug resistant gram-negative bacteria. It notes that there are currently no new drugs to treat infections from multidrug-resistant gram-negative bacilli such as Acinetobacter baumannii and Pseudomonas aeruginosa. It also discusses extended-spectrum beta-lactamases (ESBLs) which are enzymes produced by bacteria that confer resistance to many beta-lactam antibiotics. The document provides information on detecting and confirming ESBL production in bacteria.

1. DRUG RESISTANT GRAM-NEGATIVE
BACTERIA
Dr.T.V.Rao MD
DR.T.V.RAO MD 12/8/2012 1

2. NO ESKAPE ? FROM PATHOGENS
Bugs, No Drugs: No ESKAPE
– Enterococcus faecium (E), Staphylococcus aureus (S), Klebsiella
pneumoniae (K), Acinetobacter baumannii (A), Pseudomonas aeruginosa
(P), and Enterobacter spp. (E)ate-stage clinical development
pipeline remains unacceptably lean
– Some important molecules for problematic pathogens
such as MRSA
– Few novel molecules for other ESKAPE pathogens
– No new drugs for infection due to multidrug-resistant Gram-negative
bacilli (eg, A. baumannii and P. aeruginosa)
– None represent more than an incremental advance over currently
available therapies
DR.T.V.RAO MD 12/8/2012 2

3. EXTENDED-SPECTRUM Β-LACTAMASES (ESBLS):
THE FORGOTTEN (AND UNDERRATED) MDR GNB
• Most commonly identified in enterobacteriaceae
• Plasmid-mediated
• Impart decreased susceptibility to β-lactam
antimicrobials
• Often co-resistance to
aminoglycosides, fluoroquinolones
• Carbapenems are drugs of choice for invasive
infections due to ESBL-producers
DR.T.V.RAO MD 12/8/2012 3

4. WHAT ARE ESBL
• ESBLs are enzymes
capable of
hydrolysing
penicillins, broad-
spectrum
cephalosporins and
Monobactams, and
are generally derived
from TEM and SHV-
type enzymes
DR.T.V.RAO MD 12/8/2012 4

5. ALEXANDER FLEMING
NOBEL LECTURE, DECEMBER 11, 1945
“It arose simply from a fortunate occurrence which happened when I
was working on a purely academic bacteriological problem which had
nothing to do with antagonism, or moulds, or antiseptics, or
antibiotics.”
www.nobelprize.org
Google images
DR.T.V.RAO MD 12/8/2012 5

6. WHAT IS A BETA-LACTAM?
• Abx
• Penicillin
• Cephalosporin
• Monobactam
• Carbapenem
• Bactericidal Google Images
DR.T.V.RAO MD 12/8/2012 6

7. ESBLS
• Enterobacteriaceae
• Resistance to
oxyimino-
cephalosporins and
monobactams but
not cephamycins and
carbamenems
• Susceptible to
beta-lactamase
inhibitors
Oteo, et al., 2010
DR.T.V.RAO MD 12/8/2012 7

8. GENES
• SHV
• TEM
• CTX-M
• OXA
• AmpC
DR.T.V.RAO MD 12/8/2012 8

9. CLASSIFICATION
• Ambler Classification
• Molecular class A – D
•A
• Bush-Jacoby-Medeiros Classification
• Functional group 1 – 4
•2
• 2b
• 2be
Paterson and Bonomo, 2005
DR.T.V.RAO MD 12/8/2012 9

10. ESBL EVOLUTION
• Mid 1980s
• Variants of TEM and
SHV
• Breakdown 3rd
generation
cephalosporins
• Mainly in hospital
Klebsiella
• Spread world wide
DR.T.V.RAO MD 12/8/2012 10

11. WHERE ESBL ARE LOCATED
• ESBLs are often
located on
plasmids that are
transferable from
strain to strain and
between bacterial
species.
DR.T.V.RAO MD 12/8/2012 11

12. CTX-M: ESBL EPIDEMIC
•Common ESBL worldwide, often produced by
Escherichia coli
•Often causes UTI
•Now reported in US
• Healthcare associated
• Some community
•Community-based ESBL infection raise
concern for continued increases in
carbapenem use
DR.T.V.RAO MD 12/8/2012 12

13. WHY WE NEED ESBL DETECTION
• ESBL-producing Enterobacteriaceae have been
responsible for numerous outbreaks of infection
throughout the world and pose challenging infection
control issues. Clinical outcomes data indicate that
ESBLs are clinically significant and, when
detected, indicate the need for the use of appropriate
antibacterial agents.
• Unfortunately, the laboratory detection of ESBLs can be
complex and, at times, misleading.
DR.T.V.RAO MD 12/8/2012 13

14. ESBL PRODUCING BACTERIA ARE MORE
COMPLEX ?
• Antibacterial choice is often complicated by
multi-resistance. Many ESBLproducing
organisms also expressAmpC â-
lactamases and may be co-transferredwith
plasmids mediating aminoglycoside
resistance. In addition, there is an
increasing association between ESBL
production and fluoroquinolone resistance
DR.T.V.RAO MD 12/8/2012 14

15. DYNAMICS OF ANTIBIOTIC
RESISTANCE
DR.T.V.RAO MD 12/8/2012 15

16. PENICILLINS
• 1st generation
• Extended-
spectrum
• With or w/o
beta-lactamase
inhibitor
• Broad spectrum
DR.T.V.RAO MD 12/8/2012 16

17. CEPHALOSPORINS
4th
3rd
2nd
1st
Willey, et al., 2008
DR.T.V.RAO MD 12/8/2012 17

18. ESBL DYNAMICS
• Although in in vitro tests
ESBLs are inhibited by
â-lactamase inhibitors such
as clavulanic acid, the
activity of â-lactam/â-
lactamase inhibitor
combination agents is
influenced by the bacterial
inoculum, dose
administration regimen and
specific type of ESBL
present
DR.T.V.RAO MD 12/8/2012 18

19. THE FIGHT
• Beta-lactam
• Beta-lactamase
• Beta-lactamase
inhibitor
• ESBL Google Images
DR.T.V.RAO MD 12/8/2012 19

20. THE FIGHT
BETA-LACTAM
PG
N cell
O
DR.T.V.RAO MD 12/8/2012 LYSIS 20

21. THE FIGHT
BETA-LACTAMASE
PG
beta-lactamase
N cell
O
DR.T.V.RAO MD 12/8/2012 21

22. THE FIGHT
BETA-LACTAMASE
PG
O NH
OH
cell
DR.T.V.RAO MD 12/8/2012 22

23. THE FIGHT
BETA-LACTAMASE INHIBITOR
PG
beta-lactamase
N Inhibitor cell
O
DR.T.V.RAO MD 12/8/2012 23

24. THE FIGHT
BETA-LACTAMASE INHIBITOR
PG
beta-lactamase
Inhibitor
N cell
O
DR.T.V.RAO MD 12/8/2012 LYSIS 24

25. WHAT ARE EXTENDED-SPECTRUM
Β-LACTAMASES?
• ESBLs are enzymes that mediate
resistance to extended-spectrum (third
generation) cephalosporins
(e.g., ceftazidime, cefotaxime, and
ceftriaxone) and monobactams
(e.g., aztreonam) but do not affect
cephamycins (e.g., cefoxitin and cefotetan)
or Carbapenems (e.g., meropenem or
imipenem
DR.T.V.RAO MD 12/8/2012 25

26. WHY SHOULD CLINICAL LABORATORY PERSONNEL BE
CONCERNED ABOUT DETECTING THESE ENZYMES?
• The presence of an ESBL-producing organism in a clinical
infection can result in treatment failure if one of the above
classes of drugs is used. ESBLs can be difficult to detect
because they have different levels of activity against
various cephalosporins. Thus, the choice of which
antimicrobial agents to test is critical. For example, one
enzyme may actively hydrolyze ceftazidime, resulting in
ceftazidime minimum inhibitory concentrations (MICs) of
256 µg/ml, but have poor activity on cefotaxime, producing
MICs of only 4 µg/ml. If an ESBL is detected, all
penicillins, cephalosporins, and aztreonam should be
reported as resistant, even if in vitro test results indicate
DR.T.V.RAO MD 12/8/2012 26
susceptibility

27. HOW CAN CLINICAL LABORATORY
PERSONNEL CONFIRM ESBL PRODUCTION?
• NCCLS recommends performing phenotypic confirmation of
potential ESBL-producing isolates of K. pneumoniae, K.
oxytoca, or E. coli by testing both cefotaxime and
ceftazidime, alone and in combination with clavulanic acid .
Testing can be performed by the broth micro dilution method or
by disk diffusion. For MIC testing, a decrease of > 3 doubling
dilutions in an MIC for either cefotaxime or ceftazidime tested in
combination with 4 µg/ml clavulanic acid, versus its MIC when
tested alone, confirms an ESBL-producing organism. For disk
diffusion testing, a > 5 mm increase in a zone diameter for either
antimicrobial agent tested in combination with clavulanic acid
versus its zone when tested alone confirms an ESBL-producing
organism.
DR.T.V.RAO MD 12/8/2012 27

28. CLINICAL STRATEGY TO DETECT
ESBL
CTX/CLA CAZ/CLA
CTX CAZ

29. HOW SHOULD LABORATORY PERSONNEL TEST FOR
CEFOTAXIME AND CEFTAZIDIME IN COMBINATION WITH
CLAVULANIC ACID?
• NCCLS suggests making
disks by adding 10 µl of a
1000 µg/ml stock solution
of clavulanic acid to
cefotaxime and ceftazidime
disks each day of testing .
In the future, commercial
manufacturers of
antimicrobial disks may
produce disks containing
cefotaxime and ceftazidime
with clavulanic acid.
DR.T.V.RAO MD 12/8/2012 29

30. CTX-M: ESBL EPIDEMIC
•Common ESBL worldwide, often produced by
Escherichia coli
•Often causes UTI
•Now reported in US
• Healthcare associated
• Some community
•Community-based ESBL infection raise concern
for continued increases in carbapenem use
Urban, Diag Micro Infect Dis, 2010; Sjölund-Karlsson, EID, 2011
DR.T.V.RAO MD 12/8/2012 30

31. CARBAPENEM RESISTANCE
• Emerging problem in Pseudomonas
aeruginosa, Acinetobacter
baumannii, Enterobacteriaceae (CRE)
• Risk factors include ICU stay, prolonged
exposures to healthcare, indwelling
devices, antibiotic exposures
• Long-term acute care centers (LTACs)
• Severely limits treatment options
• Increased use of older, toxic agents such as
colistin
DR.T.V.RAO MD 12/8/2012 31

32. KLEBSIELLA PNEUMONIAE CARBAPENEMASES (KPCS)
• Plasmid-mediated carbapenemases
• KPC-producing strains of Klebsiella pneumonia and other
enterobacteriaceae
• KPC-2, KPC-3
• Endemicity in many locales in the US
• Hyperendemicity in NYC
• 24% of K. pneumoniae infections were due to KPCs in 2
hospitals
• Country-wide outbreak ongoing in Israel, Greece, Columbia and
others
32
*
DR.T.V.RAO MD 12/8/2012

33. KPCS (CONT)
• Might appear susceptible to imipenem or
meropenem, but with borderline MICs per 2009 CLSI
breakpoints
• Usually Ertapenem resistant
• Modified Hodge test
• Usually only susceptible to colistin, Tigecycline and
select aminoglycosides
• Easily spread in hospitals (often requires Cohorting of
staff and patients to control)
DR.T.V.RAO MD 12/8/2012 33

34. DO ISOLATES OTHER THAN K.
PNEUMONIAE, K. OXYTOCA, OR E. COLI
PRODUCE ESBL’S?
• Other isolates of
Enterobacteriaceae, such
as Salmonella species and
Proteus mirabilis, and
isolates of Pseudomonas
aeruginosa produce
ESBLs. However, at this
time, methods for screening
and phenotypic
confirmatory testing of
these isolates have not
been determined by
NCCLS.
DR.T.V.RAO MD 12/8/2012 34

35. HOW SHOULD CEPHALOSPORIN AND
PENICILLIN RESULTS BE REPORTED?
• If an isolate is confirmed as an ESBL-producer by the
NCCLS-recommended phenotypic confirmatory test
procedure, all penicillins, cephalosporins, and
aztreonam should be reported as resistant. This list
does not include the Cephamycins (cefotetan and
cefoxitin), which should be reported according to their
routine test results. If an isolate is not confirmed as an
ESBL-producer, current recommendations suggest
reporting results as for routine testing. Do not change
interpretations of penicillins, cephalosporins, and
aztreonam for isolates not confirmed as ESBL
DR.T.V.RAO MD 12/8/2012 35

36. BACTERIA NOT TO TEST FOR
ESBLS
• Acinetobacter
• Often S to clavulanate
alone
• S. maltophila +ve result
by inhibition of L-2
chromosomal b-
lactamase, ubiquitous
in the species
DR.T.V.RAO MD 12/8/2012 36

37. NEW DELHI METALLO-BETA-LACTAMASE-1
(NDM-1)
• Carbapenemases mediating broad spectrum
resistance
• Usually found in Klebsiella pneumonia, E. coli
• Initially identified in India, Pakistan, Bangladesh
• Recovered in
Australia, France, Japan, Kenya, North
America, Singapore, Taiwan, and the United
Kingdom, Australia, Canada
• Recovered in the US (Massachussetts, Illinois and
California)
DR.T.V.RAO MD 12/8/2012 37

38. MDR GNB IN LONG TERM CARE
• Quinolone resistance increasingly common in
hospitals, long-term care and in some community
settings
• B-lactam resistance established in hospitals, many
long-term care settings
• Risk factors in long-term care for resistant Gram-
negative bacilli
• Indwelling devices
• Poor functional status
• Pressure ulcers/wounds
• Antimicrobial/quinolone exposure
• Prior hospitalization
DR.T.V.RAO MD 12/8/2012 38

39. STRATEGIES TO CONTROL THE
SPREAD OF MDR GNB
• Contact
precautions/hand
hygiene
• Environment and
source control
• Antibiotic stewardship
• Enhanced infection
control measures
• Bundles
DR.T.V.RAO MD 12/8/2012 39

40. ROLE OF THE ENVIRONMENT
• Environmental sources of contamination/infection
• Increasingly recognized as sources of infection
• Particularly important with pathogens such as
Clostridium difficile, Norovirus, Acinetobacter
spp.
• Bleach preparations are more effective for some
pathogens (still need cleaning)
• Latest technology being tested: UV
light, hydrogen peroxide vapor
DR.T.V.RAO MD 12/8/2012 40

41. ENVIRONMENTAL CLEANING
• Adequacy of cleaning of patients’ rooms
suboptimal
• Improve monitoring and feedback of efficacy of
cleaning
• Direct observation and culturing not efficient, time-
consuming and expensive
• Other options: ATP bioluminescence and
fluorescent dyes
• Monitor process, efficacy of cleaning
DR.T.V.RAO MD 12/8/2012 41

42. SUPPLEMENTS TO ROUTINE ENVIRONMENTAL
CLEANING
• Disinfection units that decontaminate
environmental surfaces
• Must remove debris and dirt in order for
these units to be effective
• Two most common methods
• UV light
• Hydrogen peroxide (HP)
DR.T.V.RAO MD 12/8/2012 42

43. CHLORHEXIDINE GLUCONATE (CHG)
• Broad-spectrum
antimicrobial disinfectant
• Preferred agent for skin
preparation prior to
insertion of vascular
catheter and prior to
surgery
• Studied for “source
control”, decrease in
degree of contamination of
patients by problem
hospital pathogens
DR.T.V.RAO MD 12/8/2012 43

44. ENHANCED INFECTION CONTROL PROCESSES
• Active Surveillance
• Use of “screening” cultures to identify patients colonized with
pathogens (usually MDR) of interest
• Goal is to prevent spread in the hospital by identifying patients who
are colonized and intervening to prevent spread
• Most experience is with Gram positive pathogens
• Limited use for some pathogens (due to low sensitivity)
• Cohorting of patients
• Dedicated staff
DR.T.V.RAO MD 12/8/2012 44

45. ANTIMICROBIAL STEWARDSHIP - GOALS
• Optimize appropriate use of antimicrobials
• The right agent, dose, timing, duration, route
• Optimize clinical outcomes
• Reduce emergence of resistance
• Limit drug-related adverse events
• Minimize risk of unintentional consequences
• Help reduce antimicrobial resistance
• The combination of effective antimicrobial stewardship and
infection control has been shown to limit the emergence and
transmission of antimicrobial-resistant bacteria
Dellit TH et al. Clin Infect Dis. 2007;44(2):159–177; . Drew RH. J Manag Care Pharm.
2009;15(2 Suppl):S18–S23; Drew RH et al. Pharmacotherapy. 2009;29(5):593–607.
DR.T.V.RAO MD 12/8/2012 45

46. BUNDLES
• A bundle is a structured
way of improving the
processes of care and
patient outcomes: a
small, straightforward
set of evidence-based
practices that, when
performed collectively
and reliably, have been
proven to improve
patient outcomes.
Resar R, Joint Commission Journal
DR.T.V.RAO MD 12/8/2012 46
on Quality and Patient Safety.

47. CONCLUSIONS
• MDR GNB are growing in prevalence in multiple
geographic locales
• Occur in a variety of healthcare associated
settings
• Even in the community
• Antimicrobial stewardship is here to stay
• Problem is compounded by dry pharmaceutical
pipeline
• Novel methods to control spread of MDROs are
attractive but not clearly effective/cost-effective
DR.T.V.RAO MD 12/8/2012 47

48. NEW DELHI METALLO-BETA-LACTAMASE-1
(NDM-1)
• Carbapenemases mediating broad spectrum
resistance
• Usually found in Klebsiella pneumonia, E. coli
• Initially identified in India, Pakistan, Bangladesh
• Recovered in
Australia, France, Japan, Kenya, North
America, Singapore, Taiwan, and the United
Kingdom, Australia, Canada
• Recovered in the US (Massachussetts, Illinois and
California)
DR.T.V.RAO MD 12/8/2012 48

49. STILL THE BEST WAY TO PREVENT SPREAD OF
INFECTIONS AND DRUG RESISTANCE IS ……
DR.T.V.RAO MD 12/8/2012 49

50. VISIT ME FOR MORE ARTICLES OF INTEREST
ON INFECTIOUS DISEASES ……
DR.T.V.RAO MD 12/8/2012 50

51. • Programme Created by Dr.T.V.Rao MD
for Microbiologists in the Developing
World
• Email
• doctortvrao@gmail.com
DR.T.V.RAO MD 12/8/2012 51