results and experience of barn esbl project · results and experience of barn esbl project marina...
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Results and experience of BARN ESBL project
Marina Ivanova and project team
Tallinn
13.05.2013
Aim of the project
Improvement of detection and surveillance of resistance caused by extended spectrum beta-lactamase (ESBL) in Enterobacteriaceae in Baltic region (Estonia, Latvia, Lithuania and St Petersburg area of Russia).
Why ESBL?
1. Beta-lactams are most widely used AB group
2. Incidence is increasing rapidly (compared to MRSA, VRE etc)
Insufficient data on ESBL epidemiology of Baltic region
K. pneumoniae resistant to 3 GC – incidence is increasing
3. Many different types/genes and regional differencies
4. Rapid spread of new types
5. Colonisation of healthy people
24 of 100 travellers were colonised by ESBL pos E. coli and 5 of 21 remained colonised for longer than 6 months after the travel
6. Higher mortality and costs of treatment
Aims of the project in detail
• 1. Introduction and evaluation of phenotypic screening and confirmation algorithms of ESBLs (incl ESBL-A, ESBL-CARBA and ESBL-M)
• 2. Detection of main phenotypic and molecular markers in E. coli and K. pneumoniae strains resistant to 3rd generation cephalosporins in Baltic region
• 3. Creation of culture collection of resistant E. coli and K. pneumoniae strains for future studies
• 4. Improvement of antimicrobial resistance detection and resistance surveillance in Baltic region
• 5. Educational and scientific cooperation between EU (Baltic and Nordic states) and Russia
Project history: activities • Feb 2011 - 1st BARN WS: presentation of project
proposal
• March to Sept 2011: Estonian Pilot study (4 labs)
• Nov 2011: 2nd BARN WS: participants meeting, final protocol
• Jan - May 2012: study period
• Introduction of ESBL screening/confirmation methods
• Collection of strains and data in local labs
• Meetings in St. Petersburg and Vilnius
12
Project history: activities • June-Sept 2012:
• Collection of strains and data to study center
• ID- MALDI-TOF
• Extraction of DNA
• Master students training in Stockholm
• 2013
• ECCMID posters preparation and presentation
• Final meeting in Tallinn
• Final report to BARN
• Initiatives for involvement of new countries
• Preparation of publications
13
21 labs/hospitals
Collaboration
Strains/data collecting coordinators
Jolanta Miciuleviciene, Lithuania, Vilnius (City) University Hospital Ruta Ambrazaitiene, Vilnius University Hospital Gintaras Makstutis, Siaulai Hospital Arta Balode, Latvia, Pauls Stradins University Dace Rudzite, Riga East Clinical University hospital Tatjana Djundika, Liepaja Hospital Lidia Kaftyreva, Russia, St. Petersburg Pasteur Institute Svetlana Egorova, St. Petersburg Pasteur Institute Nataliya Vedernikova, St. Petersburg Hospital № 4 Lidia Lipskaya, St. Petersburg Hospital № 40 Olga Morozova, St. Petersburg Children's hospital № 1 Tatyana Kurchikova St. Petersburg Children's hospital № 17 Maria Paysetchkaya, St. Petersburg Children's hospital № 5 Marina Smirnova, St. Petersburg Hospital № 16 Irina Konovalenko, St. Petersburg Hospital № 31 Marina Ivanova, Estonia, ITKH Svetlana Rudenko, PERH Kaisa Kirs, LTKH Krista Lõivukene, TÜK Natalja Kamõnina, IVKH
Scientific support and activities Estonia-Sweden-Norway
Barbro Olsson Liljequist, Petra Edquist
– SmittskyddsInstitutet
Paul Naaber – Stavanger University Hospital
Paul Naaber, Epp Sepp, Siiri Kõljalg, Marina Ivanova – Tartu University
Anastasia Pavelkovich, Jana Lillo, Kristiine Pai – Master students
Outputs, results
• Optimization of lab diagnostics and detection of resistance mechanisms
• Sustainable and integrated surveillance network base for ESBLs in Baltic-Nordic region
• Scientific publications about ESBL epidemiology in Baltic region
• National guidelines (for labs and IC - isolation).
Collection of strains
970 ESBL screening positive strains (13 140 screened)
433 strains of E. coli
537 strains of K. pneumoniae January – May 2012
Clinical isolates, 1 per patient
Collection ID check
• All strains were checked with MALDI Biotyper Microflex LT (Bruker Daltonics)
• Only confirmed (EC and KP) strains were included into collection
AST and phenotypical resistance screening
• ESBL A, ESBL M, ESBL CARBA – first EC and KP isolates, meeting the criteria for ESBL phenotypical screening
– Screening (3 gen cefalosporins – ceftazidim, cefotaxim NS)
– Screening (carbapenems – ertapenem, meropenem NS)
Options for ESBL screening
Options Antibiotic(s) Methods Interpretation
(screening positive)
1. recommended Ceftazidime AND
cefotaxime
(ceftriaxone can be
used instead of
cefotaxime)
Disk diffusion or
MIC – Etest or
MIC automated
system
Any non-susceptible (i.e.
Ceftazidime or
cefotaxime or
ceftriaxone)
2. alternative VITEK or other
expert system
e.g. VITEK Non-susceptible
(ceftazidime or
cefotaxime or
ceftriaxone) or VITEK
alert for possible
ESBL/AmpC
3.Carbapenemase
screening
Ertapenem or
Meropenem or
Imipenem
Disk diffusion or
MIC
Any non-susceptible
•
Cefpodoxime < S OR
Cefotaxime and/or Ceftazidime < S (S2)
Ertapenem and/or Meropenem and/or
Imipenem MIK <S (S3)
L1 most common:
E. coli
K. pneumoniae
P. mirabilis
Salmonella spp.
Shigella spp.
L2
K. oxytoca
P. vulgaris
C. koseri
L3
Enterobacter spp.
C. freundii
M. morganii
H. alvei
Serratia spp.
Providencia spp.
ESBL test (T2) AmpC
test
(T1)
NEG or
Non-
detectabl
e
MBL, KPC, AmpC (T4)
ESBL
(R4)
POS
E. coli
Shigella
KR/PL
AmpC
(R1)
K. oxytoca
K. pneumoniae
P. mirabilis
Salmonella spp.
PL AmpC (R2)
KR
AmpC
(R3)
Ceftaz
idime
ESBL
POS
MBL
(R6)
KPC
(R7)
MBL
POS
KPC POS
AmpC NEG
KPC NEG
AmpC POS
AmpC +
porin
(R1/2/3)
KR AmpC
(R3)
Irrespective of
screening results
D-test/
Cefox
(S1)
L3
Enterobacter spp.
C. freundii
M. morganii
H. alvei
Serratia spp.
Providencia spp.
POS
ESBL test (T3)
NEG or
Non-
detectable
POS
ESBL and KR
AmpC (R5)
Comments: S1-S3: Screening tests 1-3; L1-L3: Microbial lists 1-3; T1-T4: Confirmation tests 1-4; R1-7: Lab reports to clinicians and infection control 1-7 KR AmpC: Chromosomal AmpC; PL AmpC: Plasmid AmpC
Algorithm for detection of ESBLA (= ESBL inhibited by clavulanic acid)
CTX-R or CAZ-R
E.coli, K.pneumoniae, P.mirabilis, Salmonella
spp., Shigella spp.
ESBL A-test (clavulanic acid)
Positive = ESBL A
Negative => Perform ESBL M -
test
K.oxytoca, P.vulgaris, C.koseri
Chromosomal beta-lactamase
ESBL A-test only if CAZ-R
Other Enterobacteriaceae
Often chromosomal beta-lactamase
ESBL A-test if resistant to >= 2 non-betalactams
Genotyped by PCR for CTX-M, SHV, TEM or other
Algoritm for detection of aquired (plasmid-mediated) AmpC (=ESBLM)
ESBL A-negative and CTX/CAZ-R; E.coli, K.pneumoniae, (P.mirabilis,
Salmonella spp., Shigella spp.)
ESBL M-test positive (CAZ-CLOX synergy)
E.coli and Shigella spp. : PCR to differentiate between plasmid- or
chromosomally-mediated
K.pneumoniae, P.mirabilis, Salmonella spp. do not possess chrom. AmpC, therefore they have aquired AmpC
ESBL M-test negative (no synergy)
Different mechanism (loss of porins, PBP alterations etc)
Genotyped by PCR for pAmpC; CIT and DHA are most common
Algorithm for detection of carbapenemases (=ESBLCARBA)
Meropenem I/R using DD, or MIC > 0.5 mg/L
Synergy with boronic acid but
not cloxacillin
= KPC (class A carbapenemase)
Synergy with boronic acid and
cloxacillin
= AmpC and loss of porins
Synergy only with dipicolinic acid
(DPA)
= Metallo-betalactamase (e.g.
VIM, IMP, NDM)
No synergy
= ESBL and loss of porin, or OXA-48
ROSCO test evaluation results
Diagnostic tests from Rosco
Diagnostic tests for ESBLM from Rosco
CTX30
CTX30+CLOX
CAZ30
CAZ30+CLOX
Diagnostic tests for ESBLCARBA from Rosco
Molecular analysis of the collection
– Practical training of students in SmittskyddsInstitutet
– ESBL genes detection • ESBLA (CTX-M, SHV, TEM)
• ESBLM (AmpC)
• ESBLCARBA (NDM, KPC…)
ECCMID 2013 posters presented
• Epidemiology of Extended Spectrum Beta Lactamases producing E. coli and K. pneumoniae in the Baltic Sea Region
• Differences in virulence factors of E. coli isolated from the Baltic Sea Region
• The molecular epidemiology of Extended Spectrum Beta Lactamases producing E. coli and K. pneumoniae in the Baltic Sea Region
• Detection of carbapenem non-susceptible E. coli and K. pneumoniae in the Baltic Sea Region
Further analysis of the data
• Sensitivity of different screening agents and criteria (needs additional tests and work)
• Strains without confirmed mechanisms of resistance (phenotypically and/or genes)
• MIC distributions in different ESBL classes?
• Comparison of different fenotypical methods for detecton of resistance mechanisms?