Surveillance by Cultural Methods

Active surveillance by cultural methods

J. Papaparaskevas, MD, PhD, Associate Professor, Dpt. of Microbiology, Medical School, National and Kapodistrian University of Athens [email protected] © ESCMID eLibrary by author Disclosure slide for speaker at EUCIC Local module for Infection Prevention and Control

Disclosure of speaker’s interests

(Potential) conflict of interest author

Potentially relevant company by relationships in connection with event 1

Research Grants from Wyeth-Pfizer, Merck & Co, Astra- • 2 Sponsorship or research funding Zeneca, bioMerieux and LabSupplies, through the Special eLibraryAccount for Research Grants of the National and Kapodistrian University of Athens • Fee or other (financial) payment3 • Shareholder4 Speaker Honoraria from Wyeth-Pfizer and Astra-Zeneca, • Other relationship, i.e. …5 through the Special Account for Research Grants of the ESCMID National and Kapodistrian University of Athens © Definitions

author Surveillance: monitoring of behavior, activitiesby or other changing information of individuals/organism/system Microbial surveillance: continuous and systematic collection, analysis and interpretation of microbiological data Aim of microbial surveillance: planning,eLibrary implementation and evaluation of infection control practices and/or treatment strategies

ESCMID © Definitions

Geographical level (continent/nationwide) author Data: infection burden, trends in resistance, emergenceby and spread of new resistance mechanisms Aim: guidance of healthcare policy, alerts, development and adaptation of international/national guidelines

Institutional level Data: infection rates (benchmark), eLibrarylocal trends in resistance, import and spread of new resistance mechanisms Aim: Guidance and evaluation of infection control strategies, detection of outbreaks, development and adaptation of local formulary Patient level ESCMID Data: colonization status, infection status (MDR strain) Aim: Guidance of© barrier precautions, guidance of (empirical) antibiotic therapy Microbiological cultures

author For diagnosis: √ Sampled upon clinical suspicion of infection by √ Targeted at focus of infection, ‘deep sites’, avoidance of ‘normal flora’ √ Aimed to document infection (probability and site) and to modify empirical antibiotic therapy eLibrary For surveillance: √ Sampled upon regular basis, regardless of clinical suspicion of infection √ Targeted at preferentially colonized sites, ‘superficial sites’ √ Aimed to document colonization for infection control practice and/or for anticipation of possibleESCMID infection © Which body site for which pathogen

Enterobacterales (ESBL-positive, Carbapenem-resistant,author etc.) √ The consensus dictates rectal or stool samples byto be used for detection of CRE- or ESBL-positive, and/or other multidrug-resistant Enterobacterales √ Perirectal samples are considered equal and safer for neutropenic patients √ Inguinal samples are also being considered eLibrary

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Krishna et al, 2018; Richter et al, 2017 author by

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Stier et al, 2016 Which body site for which pathogen

author Non-fermenting rods (Pseudomonas and Acinetobacter spp.) √ No consensus exists by √ Usually culture from multiple sites to increase sensitivity (sensitivity can be as low as 50% for Acinetobacter, despite multiple sampling) √ Body sites include: • Rectum • Skin eLibrary • Pharynx • Nose • Wounds • Urine • Tracheal aspirateESCMID (if patient on mechanical ventilation) ©

Krishna et al, 2018 Enterobacterales: CRE or CP-CRE?

 Carbapenem-Resistant Enterobacterales (CRE) authorare those that are non-susceptible (i.e., intermediate or resistant)by to carbapenems √ Main mechanisms include porin alterations or production of enzymes or active efflux or combinations (i.e. ESBL/AmpC + porin)  The CDC CRE definition: √ Isolates that are resistant to any carbapenem antimicrobial (i.e., MIC ≥4 μg/ml for doripenem or meropenemeLibraryor imipenem OR ≥2 μg/ml for ertapenem) OR √ Documented to produce one or more carbapenemases In addition √ For bacteriaESCMID that have intrinsic imipenem nonsusceptibility (i.e., Morganella©, Proteus, Providencia), resistance to carbapenems other than imipenem is required https://www.cdc.gov/hai/pdfs/cre/CRE-guidance-508.pdf Enterobacterales: CRE or CP-CRE?

author  Carbapenemase-Producing Carbapenem-Resistant Enterobacterales by (CP-CRE) are those that produce enzymes such as: √ Klebsiella pneumoniae carbapenemase (KPC) √ New Delhi metallo-β-lactamase (NDM) √ Verona Integron-encoded metalloeLibrary-β-lactamase (VIM) √ Oxacillinase-48-type carbapenemases (OXA-48 or equivalent) √ Imipenemase (IMP) metallo-β-lactamase

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https://www.cdc.gov/hai/pdfs/cre/CRE-guidance-508.pdf Enterobacterales: CRE or CP-CRE?

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Alizadeh et al, 2018 Enterobacterales: CRE or CP-CRE?

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Viau et al, 2016 Enterobacterales: CRE or CP-CRE?

author  Differentiating between CP-CRE and non-CP-CRE is complicated by by a number of issues √ The wide variability in the capacity of clinical and public health laboratories to perform testing √ The geographical and epidemiologicaleLibrary distribution of the mechanisms √ The clinical and epidemiological significance of the mechanisms √ The absence of consensus

 In that respectESCMID what is being done today is detection of CRE and then differentiation© of the underlying mechanism

Krishna et al, 2018 The CDC guidelines author by

√ Inoculation of rectal swabs into 5 ml of Trypticase soy broth (TSB) along with a 10 μg ertapenem or meropenem disk √ After incubation, the broth is sub-cultured to MacConkey agar, and lactose-fermenting colonies are investigated for carbapenemaseeLibraryproduction or carbapenem resistance

ESCMID √ These guidelines© (from 2013-2014) are no longer valid √ The evolution of resistance mechanisms renders guidelines obsolete very rapidly Surveillance by culture methods

 The CDC protocol author  Chromogenic media by √ CHROMagar KPC / mSuperCARBA(CHROMagar Microbiology) √ Brilliance CRE agar (Oxoid – Thermo Scientific) √ ChromID Carba Smart / Carba / OXA-48 (bioMerieux) √ Spectra CRE (Remel) √ Chromatic CRE / OXA-48eLibrary (Liofilchem) √ Other similar and/or equivalent  Non- chromogenic media, MacConkey or Drigalski agar plates: √ w ith meropenem/ertapenem disk √ with meropenemESCMIDdisks (plain, +EDTA, +boronic, +EDTA/boronic) √ supplemented© with imipenem / imipenem / ertapenem

Alizadeh et al, 2018 Surveillance by culture methods

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Viau et al, 2016 CHROMagar KPC Remel Oxoid Spectra CRE Brilliance CRE

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K. pneumoniae KPC eLibrary

ESCMID E. coli KPC © bioMerieux bioMerieux Liofilchem ChromID Carba Smart ChromID Carba Chromatic CRE ChromID OXA-48 Chromatic OXA-48 KPC/NDM OXA-48

E. coli KPC K. pneumoniae author OXA-48 by

eLibrary E. coli KPC and K. pneumoniae KPC

ESCMID © CHROMagar CHROMagar bioMerieux Oxoid ESBL CTX-M ChromID ESBL Brilliance ESBL

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eLibraryE. coli E. coli

ESCMID K. pneumoniae K. pneumoniae © Commercially available chromogenic media

author ♦ The choice (and concentration) of antimicrobial(s)by selected for inhibition of carbapenem-susceptible Enterobacterales is the most critical factor that influences sensitivity and specificity ♦ The inclusion of antimicrobials other than carbapenems may have eLibrary advantages over the inclusion of a carbapenem ♦ However, the ingredients of commercially available chromogenic media are typically undisclosed ESCMID ©

JD Perry, 2017 SUPERCARBA medium

Drigalski agar medium with: √ Ertapenem at a concentration of 0.25 μg/ml author √ ZnSO4 (zinc sulfate) at a concentration of 70 μg/ml to improveby expression of metallo-β- lactamases √ Cloxacillin at a concentration of 250 μg/ml for preventing growth of isolates expressing high levels of cephalosporinases, such as Enterobacter, Morganella, Serratia

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Nordman et al, 2012 √ The SUPERCARBA medium has the highest sensitivity (96.5%)author (detecting virtually all carbapenemase producers including OXA-48–like producers) as compared to Brilliance CRE (76.3%) and CHROMagar KPC (43%) by √ It’s specificity however was lower than CHROMagar KPC but higher than Brilliance CRE, although all media had low specificities (57% to 68%)

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Girlich et al, 2013 CHROMagar™ mSuperCARBA™

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ESCMID © CDC protocol or chromogenic media

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Disadvantages of the CDC broth method: by √ Significantly more labor-intensive than use of chromogenic media √ Longer time to detection (an extra day is required) √ Lacks information regarding likelyeLibrary species identification √ CPE may not be detected if they fail to ferment lactose on MacConkey agar

ESCMID © MacConkey agar with ertapenem disks, streaked to four quadrants for colony isolation author

√ Swabs streaked inby four quadrants for colony isolation √ 10 μg ertapenem disk placed at junctions of quadrants 1/2 and 2/3 √ Inoculation is being done either directly from the swab,eLibrary or after a 24 -48h broth inoculation step √ Colonies growing within a 27 mm radius from the disk are potentially CRE and their mechanisms need to be confirmed ESCMID ©

Lolans et al, 2010 MacConkey agar with ertapenem/imipenem/meropenem disks, streaked to four quadrants for colony isolation author by

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Optimal zone diameters are: √ <24mm (ertapenem) √ <34mm (meropenem) √ <32mm (imipenemESCMID) ©

Blackburn et al, 2013 MacConkey agar with meropenem disks (plain, +EDTA, +boronic, +EDTA/boronic), streaked for confluent growth

A difference of author5 mm in the inhibition zone between the MER disk without and with the inhibitorsby (PBA, EDTA, or both) was considered a positive result for the detection of KPC, MBL, or both carbapenemases, respectively eLibrary

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Pournaras et al, 2013 author by

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JD Perry, 2017 author by

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Richter et al, 2017 author by

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Seqyere et al, 2015 Overall comparison

√ CHROMagar KPC efficiently detects VIM and KPC but expresses poorauthor sensitivity for OXA-48 producers, which accounts for its overall lower sensitivity √ CHROMagar KPC is noted for its enhanced ability to detect highlyby resistant CREs (higher MIC) √ Isolates producing NDM-1 may be inhibited by CHROMagar KPC if the isolates have a relatively low MIC to meropenem (2 μg/ml)

√ Brilliance CRE agar is marketed for the isolation of CRE rather than CP-CRE √ Relatively low specificities have been recordedeLibrary due to its inability to inhibit certain AmpC- and/or ESBL-producing isolates √ Briliance CRE has higher sensitivity and specificity in detecting carbapenemase-producing Klebsiella pneumoniae than for other Enterobacterales ESCMID ©

Seqyere et al, 2015 Overall comparison (cont.)

√ ChromID CARBA was proven to be effective in comparative studies authorof media in countries where KPC is dominant, but had limited efficacy in countries where OXA-48 is dominant √ Pre -enrichment of rectal swabs in MacConkey broth improvedby slightly the sensitivity √ To address this, the manufacturer offers a biplate (chromID CARBA SMART) that combines two media in a single petri dish: chromID CARBA and chromID OXA-48.

√ The SUPERCARBA is the only medium able to detect all carbapenemases types (A, B, OXA-48) √ The stability of the medium however is limitedeLibrary to 1 week √ Its commercially available adaptation, CHROMagar mSuperCarba, has an improved shelf life of 1 month

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Seqyere et al, 2015 Enterobacterales: what about colistin?

author  Colistin resistance among Gram(-) is the newby endeavor of the chromogenic media surveillance and diagnosis procedure

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√ Eosin methylene blue (EMB) eLibraryagar supplemented with polymixin, daptomycin and amphotericin B √ Not a chromogenic medium √ Directly from swabs/stool without incubation period ESCMID © √ Overall, specificity was 100.0% [95% CI: 97.8–100.0] and author sensitivity 88.1% [95% CI: 77.5–94.1] √ False negatives corresponded to three Enterobacter spp. by (MIC>64 mg/L), two Salmonella spp. (MIC=16 mg/L), one E. coli (MIC=4 mg/L), and one K. pneumoniae (MIC=8 mg/L) √ CHROMID Colistin R agar appears to be a sensitive and specific chromogenic agar for screening colistin-resistant Enterobacterales, including those carrying mcreLibrary-1 gene

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Garcia-Fernandez et al, 2019 author by

√ Directly from swabs/stool without pre-incubation period √ But, no sensitivity or specificity data are available eLibrary

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Abdul Momin et al, 2017 author by

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author ♦ Chromogenic media have clear advantages over bythe CDC method or the use of MacConkey agars supplemented with a carbapenem or used in conjunction with carbapenem disks

♦ It is especially difficult to establish which, if any, chromogenic medium is optimal for detection of CPE dueeLibrary to the different types of carbapenemase that may be encountered and the dominance of particular types in certain geographical regions

Viau et al, 2016 author by A detailed statistical meta-analysis of published studies concludes that:

√ ChromID CARBA and the non-chromogenic SuperCarba medium have excellent sensitivities for class A -lactamases (e.g., KPC) that rival that of real-time PCR

√ The CDC broth method was generally eLibraryinferior to chromogenic media

√ For other media, and other carbapenemase types, there was insufficient evidence to draw firm conclusions

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Viau et al, 2016 Surveillance by cultural methods

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Viau et al, 2016 And what about non-fermenters?

√ No consensus exists regarding the screening body siteauthor

√ Media specific for Pseudomonas aeruginosa areby the CHROMagar™ Pseudomonas, the CHROMID® P. aeruginosa Agar, the Chromogenic Pseudomonas Agar and other equivalent √ Media specific for Acinetobacter spp. are the CHROMagar™ Acinetobacter, the ChromogeniceLibrary Acinetobacter Agar and other equivalent √ Nevertheless, all these are focused for species detection and not on resistance detection √ There are many adaptations of these media using different antibiotic concentrations, but the results are controversial ESCMID √ Some of the© media used for Enterobacterales are also capable to screen for Pseudomonas aeruginosa and/or Acinetobacter spp. Factors to consider when implementing a screening program based on cultures

♦ Epidemiology of resistance in the institution author √ Prevalence of each antibiotic resistance mechanismby √ Ability to identify high-risk groups ♦ Availability and cost of isolation beds ♦ Existing logistics for collecting specimens ♦ Current clinical microbiology laboratory capabilities √ Availability of molecular diagnosticeLibrary tools √ Available technologists and ability to accommodate testing √ Experience developing and validating in-house assays √ Experience/availability of other technologies to detect resistance like Carba NP or Blue-Carba, inhibition disk assays (disk synergy test), UV spectrometryESCMID or MALDI-TOF MS, Modified Hodge test, etc. ©

Viau et al, 2016 Thank you for author your attention by

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