Invited lecture, Communicable disease control module, Master of Public Health (MPH) course, University of Sheffield, May 2017

1. Carbapenem–resistant gram-negative
bacterial infections:
Global epidemiology, challenges and threats
Evangelos Kritsotakis
Lecturer in Epidemiology & Medical Statistics
School of Health & Related Research
University of Sheffield
e.kritsotakis@sheffield.ac.uk
May 2017

2. • Basic concepts/issues: HAIs, Antibiotics, AMR
• Global Epi: features and current situation in the spread of CR-GNB,
• Challenges and threats: the example of the VIM and KPC epidemics
in Greece and the KPC epidemic in Italy,
• Risk factors for of CR-GNB infections,
• Clinical impact of CR-GNB infections,
• Treatment options for CR-GNB infections.
Overview
Overview

3. Basic
concepts & issues

4. Healthcare-associated infections (HAIs)
Healthcare-associated infections (HAIs)
• HAIs develop either as a direct result of healthcare interventions such as medical or surgical
treatment, or from being in contact with a healthcare setting.
• Point Prevalence, per 100 inpatients in acute-care hospitals:
USA 2011: 4% (95%CI: 3.7 – 4.4)
EU/EEA 2012: 6% (95%CI: 5.7 – 6.3)
Developing countries: 11% (95%CI: 8.1 – 13.9) , high quality studies: 13.5%
• Incidence, number of patients acquiring at least one HAI, per year:
USA : 648 000 (95%CI: 246 400 - 987 300)
EU/EEA: 3 200 021 (95%CI: 1 948 862 – 5 234 253)
ECDC PPS Surveillance Report 2013,
Magill S. et al NEJM 2014
Allegranzi et al Lancet 2011

5. Burden of HAIs on European population health
Burden of HAIs on European population health
DALYs = number of years lost due to ill-health, disability or early death.
PLOS Medicine 13(10): e1002150. https://doi.org/10.1371/journal.pmed.1002150
YLD = years lived with disability
YLL= years of life lost (to pre-mature mortality)
Total DALYs = 501 per 100,000 general population

6. Burden of HAIs in Europe (EU/EEA)
Burden of HAIs in Europe (EU/EEA)
DALYs = number of years lost due to ill-health, disability or early death.
Disease DALYs
per 100,000 population
HAIs (6 main types) 501
Influenza 71
TB 53
HIV infection 48
Sources:
ECDC study, 2011-2012 data, PLOS Medicine 13(10): e100215
ECDC/BCoDE study, 2009-2013 data, European Journal of Public Health 25(suppl 3); Oct 2015

7. Antibiotics
Antibiotics

8. Timeline: discovery of antibiotics
Timeline: discovery of antibiotics

9. Antimicrobial Resistance (AMR)
Antimicrobial Resistance (AMR)
• How does AMR emerge in bacteria?
- by naturally occurring pre-existing resistance mechanisms
- by de novo gene mutations (spontaneous and random)
- by horizontal transfer of mobile genetic elements.
• Protective mechanisms include:
- preventing entry of the antibiotic
- exporting the antibiotic
- producing enzymes that destroy or modify the antibiotic
- making changes to the antimicrobial target.
Holmes et al., Lancet 2016; 387: 176–87
Beaber et al., Nature 2004; 427: 72–74.

10. Antimicrobial Resistance (AMR)
Antimicrobial Resistance (AMR)
• How may antibiotics promote AMR ?
- by exerting selective pressure
- by facilitating transmission of resistant bacteria
- by facilitating transmission of antibiotic resistance genes
Holmes et al., Lancet 2016; 387: 176–87
Beaber et al., Nature 2004; 427: 72–74.
selective pressure

11. Nathan S. McClure, and Troy Day Proc. R. Soc. B
2014;281:20141861
Timeline of antibiotics against subsequent evolution of resistance
Timeline of antibiotics against subsequent evolution of resistance

12. Advanced Drug Delivery Reviews, Volume 78, 2014, 3–13
http://dx.doi.org/10.1016/j.addr.2014.08.003
Timeline of antibiotic discovery against the development of AMR
Timeline of antibiotic discovery against the development of AMR

13. Carbapenem resistance:
Global
Epidemiology

14.  Klebsiella pneumoniae
(and other enterobacteria, e.g. E.coli)
 Pseudomonas aeruginosa
 Acinetobacter spp.
Important Gram-negative nocomial
pathogens with carbapenem resistance issues

15. Microbiology of HAI
ECDC HAI PPS 2012
Source: ECDC PPS surveillance report, 2013
% isolation in healthcare-associated infections
ECDC Point Prevalence Study
231459 patients (15000 HAIs)
947 hospitals
33 countries

16. • Carbapenemases are enzymes that hydrolyse (destroy)
carbapenem antibiotics making the organism resistant
• Carbapenems: meropenem, imipenem, ertapenem,
doripenem (all IV)
• These antibiotics are the ‘last line of defence’ as they are the
broadest spectrum/most effective available
• Carbapenemase-producers are usually resistant to most other
antibiotics too.
Carbapenemases & Carbapenems
Carbapenemases & Carbapenems

17. Ambler Class B
Metallo-enzymes (MBL)
VIM (Pseudomonas, Klebsiella)
NDM (Klebsiella)
IMI, SPM, GIM, SIM
Ambler Class D
Oxacillinases
OXA-48 (Acinetobacter)
Not only genetic and biochemical classification, but also
Different epidemiology
Differences in profiles of resistance
Carbapenemases
Carbapenemases
Ambler Class A
Serine
KPC (Klebsiella)
IMI (Enterobacter)
GES (Pseudomonas)
SME (Serratia)

18. Resistance profiles
Resistance profiles
Nordmann P, Médecine et maladies infectieuses 2014; 44: 51–56

19. Global spread
Global spread
 KPC is nowdays endemic in the USA, Greece, Italy, Israel,
China and Latin America
 VIM was endemic in Greece (but KPC has taken over now)
and has substantial spread in Italy.
 NDM primarily spread in the Indian subcontinent, but
sporadic cases have been reported everywhere in Europe
 OXA have spread mainly around the eastern and southern
parts of the Mediterranean basin.

20. Munoz-Price LS et al. Lancet Infect Dis 2013; 13: 785–96
KPC Global spread
KPC Global spread

21. NDM Global spread
NDM Global spread
Front. Microbiol., 13 June 2016 |
https://doi.org/10.3389/fmicb.2016.00895

22. Carbapenem-NS isolates from the EARS-NET, 2015
K. pneumoniae
P. aeruginosa
Acinetobacter spp.
E. coli
Source: http://ecdc.europa.eu/en/publications/Publications/antimicrobial-resistance-europe-2015.pdf

23. ECDC HAI PPS 2012
Antimicrobial resistance markers
for healthcare-associated infections
*Source: ECDC, 2012 (results as of 23/11/2012)

24. Challenges and Threats:
Examples from
Greece and Italy

25. 2002:
VIM-1 in 17 K.pneumoniae isolates in 3 ICUs in Athens
“VIM-type genes have been spread via transferable plasmids in
the enterobacteria of the hospital flora in Greece”.
Giakkoupi P et al. JCM 2003; 41:3893-6
2001:
Isolation of E.coli with blaVIM-1 gene, hospital in Piraeus
Miriagou V et al. AAC 2003; 47:395-397.
Four sporadic VIM-1 E.coli isolates in Un. hospital, Crete
Scoulica E et al. DMID 2004; 48:167-172.
Class I integron with different structures suggesting a different evolution process
rather than a transfer and spread of the mobile element between the hospitals.
The Resistance spread
Inter-species resistance spread
Inter-species resistance spread

26. 2003: MBL production sporadically in E. cloacae in
Galani et al JAC 2005;55:634-8
2004-5: Outbreak of 7 MBL producing P. mirabilis in a
hospital in Thessaloniki, Vourli et al. CMI 2006; 12:691-4.
3 isolates, E.coli (2002), E. aerogenes (2003) & P. mirabilis (2004) producing
MBL VIM-1 encoded by same conjugative plasmid. Galani et al JAC 2007; 53 578-9
M. morganii (2005), Serratia liquefaciens, Klebsiella oxytoca (2006),
Providencia stuartii (2007) EID 2006:981–3, IJAA 2008:540-1, JAC 2007:183–4
Epidemic due to successful horizontal transfer of genetic material
between species.
May also facilitate their propagation into the community.
VIM-1 in Enterobacteriaceae
VIM-1 in Enterobacteriaceae

27. Spread of MBL-producers in the Community
Spread of MBL-producers in the Community
2005-8, Serres General Hospital:
45 patients with UTI or bacteremia
caused by VIM-2 P.aer, 40 reported
previous hospitalization 1mo-1y
before, 5 without link to health-
care facilities
2007-8, Serres General Hospital:
12 patients with UTI caused by VIM-
1 K.pn, 2-4 months after discharge.
Only 2 patients with known
colonization during recent
hospitalization
2005-7, Serres General Hospital:
9 outpatients with UTI cause by K.
oxytoca. All patients had been
hospitalized or exposed to health
care facilities during the preceding
year.

28. Eurosurveillance 2008; 13:1-3
CR K.pneumoniae (VIM-1) in
• 3 Hospitals in 2002
• 29/40 Hospitals in 2007
Establishment of endemicity
By 2007 VIM-1 MBL enzyme was responsible for the extremely high C- resistance
rates in K.pneumoniae in Greece: 75% in ICUs and 30% in med & surg wards

29. 2007–2008: Outbreaks of infection and/or colonization
due to KPC-2 producing K.pneumoniae identified in
hospitals in
• Crete (May 2007)
Maltezou et al. J Infect 2009; 58:213–9
• Athens (September 2007)
Souli et al. Clinical Infectious Diseases 2010; 50:364–73
• Thessaloniki (October 2007)
Pournaras et al. Antimicrob Chemother 2009; 64:348–52
The KPC spread
The KPC spread

30. Giakkoupi P et al. JAC 2011; 66:1510

31. Percentage (%) of IMIPENEM-R K.pneumoniae from all
specimens of Greek hospitals 1998-2012
www.mednet.gr/whonet
VIM
KPC

32. Bloodstream Infections due to
Carbapenem-R gram neg. pathogens
Procrustes Surveillance System 2013
Kontopidou F. Fighting AntiMicrobial Resistance Meeting, Rome, Dec. 2014
Mechanism of resistance among
Carbapenem-R Klebsiella isolates

33. Proportion
%
Year
EARS-NET database
Carbapenem-NS Klebsiella pneumoniae in Greece:
contemporary spread of two resistance mechanisms
Carbapenem-NS Klebsiella pneumoniae in Greece:
contemporary spread of two resistance mechanisms
VIM epidemic
polyclonal spread & transferable plasmids
Vatopoulos, Eurosurveillance 2008; 13(4)
Giakoupi et al Euro Surveillance 2009;14(21)
Giakkoupi et al. J Antimicrob Chemother 2011; 66: 1510–1513
0
10
20
30
40
50
60
70
80
2005 2006 2007 2008 2009 2010 2011
KPC-2 epidemic
spread mainly monoclonally

34. Intercontinental spread of KPCs
The first case of KPC outside the U.S. occurred in France,
KPC later spread in, Israel, Greece, China, and Colombia
KPC-3
KPC(+) K.pn NC, USA 1996
Until 2005, the geographical distribution of
KPC-2 & KPC-3 in K. Pneumoniae
was limited to the Eastern United States
KPC(+) K.pn
Tel Aviv, Israel 2006
Yigit et al. AAC 2003
Navon-Venezia et al. AAC 2009
KPC-2

35. 0
10
20
30
40
50
60
70
80
2005 2006 2007 2008 2009 2010 2011
Proportion
%
Year
Carbapenem-NS Klebsiella pneumoniae in Greece and Italy
Carbapenem-NS Klebsiella pneumoniae in Greece and Italy
Greece
Italy
KPC-2 epidemic
VIM epidemic
KPC-3 epidemic
EARS-NET database

36. Giani et al – JCM 2009
Fontana et al – BMC Res Notes 2010
Marchese et al – J Chemother 2010
Ambretti et al – New Microb 2010
Gaibani et al – Eurosurv 2011
Mezzatesta et al – CMI 2011
Agodi et al – JCM 2011
Richter et al – JCM 2011
Di Carlo et al – BMC Gastroenterol 2011
late 2008
late 2008
The first reported
cases of KPC-Kp
(ST258)
KPC-producing K. pneumoniae - Italy
KPC-producing K. pneumoniae - Italy
early 2011
early 2011
AMCLI – CoSA CRE network
Frasson et al – JCM 2012
ST258, ST512 (CC258) ST512
ST258
ST101
ST15
ST147-like
late 2012
late 2012
Source: Rossolini M., 2nd ARHAI Networks Meeting, Berlin 2012

37. The ongoing challenges

38. Diversity of acquired b-lactamases amongst K.pneumoniae in Greek hospitals
Papagiannitsis et al IJAA 2012 39 178-80
256 KP isolates in 8 Hospitals,
July 2010
22 different β-lactamase
combinations
43% KPC-producers,
11% VIM-producers
Most prevalent combination :
KPC-2 + SHV-12, 33%

39. Report from New York City of 2
infections caused by "pan
resistant" K. pneumoniae
 1 patient died
 1 had continuing asymptomatic
bacteruria
CID 2009; 49: 271-274

40. PDR: resistant to all classes of antibiotics used for empirical treatment, but
not to Tigecycline (newly introduced at the time)
2 years surveillance, overall mortality 24%
Colistin reg.: mortality = 30%, discharge = 94 ± 62 days
Tigecycline: mortality = 0% , discharge = 35 ± 17 days
Conclusion:
Tigecycline appears to be effective for the successful
treatment of PDR infections, when active in vitro

41. Co-resistance in Carbapenem-R
A.baumannii, P.aeruginosa, K.pneumoniae
blood isolates in Greek hospitals
Procrustes Surveillance System 2013
Kontopidou F. Fighting AntiMicrobial Resistance Meeting, Rome, Dec. 2014

42. KP combined resistance %, EARS-NET, 2015

43. Risk factors

44. • Patient and healthcare- related
 prolonged hospital stay,
 poor functional status, severe illness, ICU stay,
 presence of invasive devices, surgery
 exposure to antibiotics
• Setting / environment -related
 status of other patients / colonisation pressure
 availability of isolation rooms
 staff-to-patient ratio
 hand-hygiene compliance
 climate / temperature / moisture (?)
Temkin et al. Ann. N.Y. Acad. Sci. 2014; 1323: 22–42
Risk factors for hospital-acquired carriage or
infection with CRE
Risk factors for hospital-acquired carriage or
infection with CRE

45. • Sought to elucidate the role of various antibiotics as risk factors for
carbapenem-R ESBL-producing K. pneumoniae,
• Improving upon several methodological shortcomings:
– Appropriate control group (patients potentially at risk for CRKP)
– Case + case – control study
– Case groups with similar co-resistance profiles
– Examine several classes of antibiotics
– Account for duration of antibiotic use
– Allow for sufficiently long exposure period
– Examine potential interaction effects between antibiotics
– Adjustment for confounding factors

46. (1) Duration of exposure to β-lactam/β-
lactamase inhibitor combinations
showed a positive relationship with
ESBL-CRKP infection risk (adjusted
OR = 1.15 per day increase; 95%CI:
1.05-1.26, P = 0.001).
(2) Carbapenems and fluoroquinolones
presented a significant interaction
effect, such that increased exposure
to fluoroquinolones amplified the
effect of carbapenems on ESBL-CRKP
infection risk.
Multivariable analysis (accounting for effects of non-antibiotic exposures
and adjusting for other antibiotics) showed that :

47. Consumption of antibiotics in the hospital sector EU/EEA countries,
ESAC- Net surveillance data 2015

48. For every 5oC increase in temperature, an increase of
3% in gram-negative bacteria was observed
(IRR=1.03; 95%CI 1.02 - 1.04)

49. Clinical impact

50. Mortality attributable to CAR-Res. K.pneumoniae infections
Mortality attributable to CAR-Res. K.pneumoniae infections
Example Type of study Outcome Mortality
Patel, ICHE 2008,
USA
Matched case-control
CRKP vs CSKP
In-hospital death,
various infections
Crude mortality: 38%
Attributable mortality: 26%
Schwaber, AAC
2008, Israel
Case-case-control,
CRKP, CSKP, NoKP
In-hospital death,
various infections
Crude mortality: 44%
Attributable (vs CSKP): 31%
Attributable (vs S-KP): 42%
Borer, ICHE 2009,
Israel
Matched case-control
CRKP vs CSKP
In-hospital death,
following BSI
Crude mortality: 72%
Attributable mortality: 50%
Chang, JMII,
2011, Taiwan
Matched case-control
CRKP vs CSKP
In-hospital death,
following BSI
Crude mortality: 94%
Attributable mortality: 44%
Ben-David, CMI
2012, Israel
Nested case-control,
CRKP, ESBL-KP, S-kp
In-hospital death,
following BSI
Crude mortality: 69%
Attributable (vs ESBLKP): 30%
Attributable (vs S-KP): 45%
Excess mortality ranging 25% - 50% (CRKP – CSKP difference)
Crude mortality in CRKP infections: 38% - 94%

51. Mortality attributable to CRE infections
Mortality attributable to CRE infections
Falagas et al EID 2014; 20:1170–5

52.  Advanced Age
 Severity of illness, severe sepsis/shock
 Severe comorbidities
 Carbapenem Resistance
 Inappropriate therapy (no active antibiotic)
 Monotherapy (???)
Ben David D et al. Clin. Microbiol. Infect 2012
Zarkotou O et al. Clin. Microbiol. Infect 2011
Mouloudi E et al. Infect. Control Hosp. Epidemiol 2010
Qureshi ZA et al. Antimicrob Agents Chemother. 2012
Daikos GL et al. Antimicrob. Agents Chemother 2014
Predictors of death due to CRKP bacteremia
Predictors of death due to CRKP bacteremia

53. Epidemiological & clinical importance of CRE
Epidemiological & clinical importance of CRE
- Potential for rapid spread in health-care settings
- Resistance is highly transmissible and fast evolving
- Between patients
- Between bacteria - plasmids
- Potential of spread into the community
- Limited/suboptimal treatment options
- XDR/PDR phenotypes
- difficulties in laboratory detection
- could take decades for new effective antibiotics
- High morbidity and mortality burden

54. Treatment options
& their effectiveness

55. Treatment options for infections with CR-GNB
Treatment options for infections with CR-GNB
- Colistin (polymyxin E)
- Commonly used, as monotherapy or as the base of combination therapy (which is better?)
- Frequent adjuvants in combination therapy: carbapenems, aminoglycosides, tigecycline
(which is better?)
- Ideal dose for colistin is uncertain.
- Polymyxin B
- Limited clinical experience, a few small case series.
- Carbapenems
- Used succesfully as adjuvants in combination regimens (for relatively low MICs, 4-8 mg/L).
- Is there a potential additive or synergistic activity with col, tig or gentamycin?
- Are carbapenem-containing combinations the best option?
- Could double-carbapenem combinations be more effective (extend to higher MICS)?

56. - Tigecycline
- Used as an adjuvant in combination therapy
- Monotherapy is not recommended
- Could increasing the dose improve patient outcomes? BMC Anesthesiol 2013;13:1-8.
- Might a triple combination regimen colistin + meropenem + tigecycline be best?
- Fosfomycin
- Limited clinical experience.
- Not available in many countries, has been used succesfully in Greece mostly as an adjuvant.
- Could it be selected as salvage therapy?
- In development
- 3-4 new drugs are expexted to be available in the near future, but none active against the
entire spectrum of CR-GNB.

57. Clin Microbiol Infect 2010; 16: 102–111
“Limited clinical experience and several in vitro synergy studies seem to support
the view that antibiotic combinations should be preferred to monotherapies.”
But, in light of the data available to date, it is currently impossible to quantify the
real advantage of drug combinations in the treatment of these infections.

58. Tzouvelekis et al. Clin Microbiol Infect. 2014 Sep;20(9):862-72
Review of 20 clinical studies:
907 serious infections with KP producing KPC (75%), VIM(21%), OXA-48 (4%)
affecting mostly ICU patients (70%)
combination therapy with two or more in vitro active agents was superior to
monotherapy, providing a clear survival benefit (mortality rate, 27.4% vs.
38.7%; p <0.001).”
“The lowest mortality rate (18.8%) was observed in patients treated with
carbapenem-containing combinations”

59. Tzouvelekis et al. Clin Microbiol Infect. 2014 Sep;20(9):862-72
Regimen B vs. regimen A: p, not significant.
Regimens C, C1 and C2 vs. regimen B: p 0.001, p 0.034, and p <0.0001, respectively.
Numbers above columns indicate the number of patients

60. Limitations of (many of) these studies
Limitations of (many of) these studies
- Inclusion of Carbapenem-susceptible isolates
(bias in favour of combination therapy)
- Small sample size
 forbids adjustment for (the many) other mortality risk factors
 many different regimens involved  more risk of chance findings
- Important confounders ignored
(appropriateness of empirical therapy and time to covering antibiotic treatment)
- Varying definitions of combination therapy
(any 2 drugs, at least one covering, two covering)
- Colistin loading doses not reported
(bias against colistin monotherapy)
- Single hospital studies
- Previous reviews lacked a rigorous meta-analysis

62. Paul et al. J. Antimicrob. Chemother. 2014;
jac.dku168
All-cause mortality for colistin
monotherapy versus combination
therapy including colistin.

64. VAP due to XDR Acinetobacter baumannii, ICU patients (n=93)
University Hospital of Heraklion, Greece (Oct.2012- Mar.2015)

65. Conclusions
Conclusions
 CR-GNB present a major challenge for health-care systems worldwide
 Can rapidly spread in health-care settings
 Resistance is highly transmissible and fast evolving,
between patients and between bacteria
 High potential of spread into the community
 Lack of new treatment options, high mortality burden
 There are weak evidence suggesting that combination therapy may be
considered the optimal therapy for CR-GNB infections, but:
 Before adopting combination therapy as a standard of care, clinicians
should judiciously examine the available data and counterweight
against the increased risk of adverse effects, such as the development,
or promotion of resistance.