Oral Presentations - Session L

Objectives: Rapid detection of carbapenemase-producing organisms (CPO) is important. Data suggest direct-from-specimen NAAT is more sensitive than culture. We evaluated three multiplex carbapenemase NAAT targeting KPC/NDM/OXA48-like/ VIM/IMP genes [BDMax Checkpoints CPO (BD), Allplex Entero-DR (Seegene), and Easyplex® SuperBug CRE Assay Version C (Amplex Diagnostics)] using 150 well-characterized (phenotypic and PCR/sequencing) GNB. Methods: 150 GNB including 122 CPO (116 targeted-CPO: 88 KPC/8 NDM/5 OXA/5 NDM+OXA/7 VIM/3 IMP; 6 non-targeted CPO: 1 GES/4 SME/1 NMC), and 28 non-CPO were tested comprising 145 Enterobacteriacaeae/5 non-Enterobactericeae GNB.  Isolates were recovered from -80^oC under ertapenem-selective pressure. Limit of detection (LOD) was calculated in triplicate using four QC strains following manufacturer direct-from-specimen-protocols using 10E4/10E5/10E6/10E7/10E8 cfu/L concentrations in ESwab transport medium with results from Xpert® Carba-R (Cepheid) as reference. Colony counts confirmed concentrations and average LOD was calculated. Accuracy was determined using ≥10-fold-higher concentrations than calculated LODs. Discrepancies were repeated. Results: LOD (cfu/L) are shown (Figure). The most sensitive to least sensitive assay was Seegene, BD, Cepheid, then Amplex. All 34 non-targeted-CPO/non-CPO were negative by all assays. 1 KPC/1 NDM/3 KPC were initially missed by BD/Seegene/Amplex, respectively but were positive on repeat testing of fresh subcultures suggesting initial lost plasmids. 2 IMP were reproducibly missed by BD and Amplex. Final CPO-detection sensitivities/specificities were 100% for all non-IMP targets; respective 95%CI were: KPC 95.0-100/87.6-100; NDM 73.4-100/95.9-100; OXA48-like 68.0-100/96.0-100; VIM 59.6-100/96.1-100. Sensitivities/specificities for IMP were 100%(38.2-100)/100%(96.2-100)(Seegene) and 33%(5.6-79.8)/100%(96.2-100)(BD&Amplex).

Conclusions: All three assays were highly-accurate (100% sensitive/specific) for detection for KPC, NDM, OXA48-like and VIM CPO.  IMP was more challenging for BD and Amplex. LOD was variable but not substantially different between assays. These results along with workflow, turn-around-time, footprint, interfaceability, cost, and laboratory needs can be used to determine suitability for different laboratories.

Objective: Carbapenemase producing enterobacteriaceae (CPE) are emerging around the world, including Canada, and are associated with case fatality rates as high as 50%.   Gastrointestional carriage of CPE may serve as the reservoir for cross-contamination in the healthcare setting, thus active surveillance is important for effective containment and outbreak prevention.   In this study we evaluate commercially available screening agars for the detection of CPE using a panel of CPE, nonCPE-carbapenem resistant and carbapenem susceptible strains. Methods: A panel of strains was assembled including clinical strains from our laboratory and highly characterized strains selected from the CDC-FDA Antibiotic Resistance Isolate Bank as indicated in Table 1.

Table 1. Carbapenemase Types in the test Panel

Two commercial agars for detection of carbapenemase producing enterobacteriaceae were compared including CHROMID^TM Carba Smart (CCS) and Colorex™ SuperCARBA (CSC).  Two different inocula were used: 10⁵CFU (high) and 10²CFU (low).   All media were incubated in accordance with the manufacturer’s recommendations.  Strain viability was confirmed by concurrently planting dilutions to a non-selective blood agar plate and carbapenemase production was confirmed present or absent from all strains by PCR. Results Sensitivities for the high inocula were 89% and 93% while the low inocula was 40% and 75% for CCS and CSC respectively.   The specificities were 75% and 59% for CCS and CSC respectively.  Both plates failed to recover four strains including K. pneumoniae IMP-4, K. oxytoca KPC-3, P. mirabilis KPC-6 and P. mirabilis KPC-2.  Interestingly, CCS failed to recover two OXA-48 like strains, including K. pneumoniae OXA-232 and K. pneumoniae OXA-181, even at the high inocula. Conclusions: CSC displayed better sensitivity, at high inocula and particularly at low inocula. Although CCS had higher specificity, overall, CSC had superior performance.

Activity of ceftobiprole against Canadian bacterial pathogens from the CANWARD study. Objectives: Ceftobiprole is a recently re-released 5^th generation cephalosporin, currently available on the Canadian and European markets. It demonstrates in vitro activity against Staphylococcus aureus (methicillin-susceptible [MSSA] and methicillin-resistant [MRSA] isolates), Streptococcus pneumoniae, ESBL-negative Enterobacteriaceae, and Pseudomonas aeruginosa. It also has improved activity against AmpC-positive Enterobacteriaceae relative to ceftriaxone and ceftazidime. Related to its broad spectrum of activity, ceftobiprole may offer a monotherapeutic option for the treatment of complicated skin and soft tissue infections and community-acquired and nosocomial pneumonia. The purpose of this study was to evaluate the in vitro activity of ceftobiprole against a contemporary collection of isolates from the CANWARD study. Methods: Isolates were collected from the ongoing CANWARD study between 2008-2010 and 2015-2017. Antimicrobial susceptibility testing was performed using broth microdilution panels following CLSI recommendations (M07, 11^th edition). Minimum inhibitory concentrations were interpreted using Health Canada breakpoints (ceftobiprole) or CLSI breakpoints (ceftazidime comparator). Where no breakpoints were available for ceftobiprole, the pharmacokinetic/pharmacodynamic breakpoint of 4mg/L was used. Results:

Conclusion: Ceftobiprole is active in vitro against S. aureus (MRSA and MSSA) and Enterobacteriaceae. It is more active in vitro than ceftazidime against Acinetobacter baumannii and species with chromosomal AmpC beta-lactamases while it is less active against Klebsiella oxytoca/Raoultella sp., suggestive that its chromosomal β-lactamase (OXY) is more active against ceftopiprole than ceftazidime. While ceftobiprole appears to have anti-Pseudomonas activity, no Pseudomonas-specific breakpoints exist and PK/PD breakpoints were derived using only one dosing recommendation (500mg IV q8h). Further studies in ceftobiprole dose, interval or infusion time may reveal that a higher species-specific breakpoint for Pseudomonas could be considered, concurrent with alternative dosing recommendations. Activity is poor against Enterococcus faecium and Stenotrophomonas maltophilia.

Objective: Early targeted antimicrobial treatment can effectively reduce the mortality rate caused by bloodstream infections (BSI) and also enhance antimicrobial stewardship efforts in reducing the use of broad spectrum antibiotics. In addition to the Vitek MS® direct from blood culture bacterial identification currently practiced at our center, the ability to perform direct susceptibility testing using the serum separator tubes (SST) has the potential to significantly reduce current turnaround time (TAT) for susceptibility results from positive blood cultures. Method:  Positive monomicrobial BACT/ALERT® blood culture bottles received in the microbiology lab between October 3 and December 31, 2018 were included. Ten milliliters of broth was aspirated from a positive blood culture bottle and injected into two 5 ml SST. The SSTs were centrifuged at 3900 RPM for 5 minutes. The supernatant was discarded and a small amount of the remaining pellet containing the bacteria was used to make a suspension equivalent to 0.5 McFarland in 0.45 % saline. Vitek 2® Susceptibility results from the pellet were compared to those performed routinely on isolates after 18hr incubation on agar plates.                                                                     Results: 100 positive blood cultures including common Gram negative and positive organisms were evaluated. All reportable antibiotics had 100% essential and categorical agreement. There was one very major error for TMP-SMX for S. aureus and 70% essential agreement but 100% categorical agreement on Nitrofurantoin for K. pneumoniae. Overall essential and categorical agreements were above the acceptable value of 90% with major and minor errors below acceptable value of 10%. Conclusions: Based on the results of this pilot, direct from blood culture bacterial susceptibility testing using the SST provides accurate results for all of the reportable antimicrobials and significantly decreases TAT. 

Background: MTOP is a fecal microbiota transplantation program.  Only healthy stool donors with no personal/family history of chronic illness are accepted.  Negative microbiology screens including stool antimicrobial-resistant organism (ARO) cultures are required.  Donors are routinely monitored and excluded if their health changes or they travel. Repeat screening is completed prior to release of donations. One donor was unexpectedly positive on repeat screening for an ESBL-producing Klebsiella pneumoniae. The purpose of this study was to determine the rate of ARO carriage among healthy stool donors. Methods: All active MTOP stool donors between March 2017 and August 2018 were included.  Stool aliquots stored at -80C from each donation were thawed and tested for ARO [methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci (VRE), ESBL, and carbapenemase-producing organisms (CPO)] by planting onto Oxoid Denim Blue agar, Oxoid Brilliance VRE agar, and Oxoid MacConkey/cefpodixime agar, respectively following clinical laboratory operating procedures. Results: Four donors actively donated during the study period.  The duration of time each participated varied from 3 months (Donors 1 and 2) to 13-14 months (Donor 3 and 4, respectively).  Eighty donations were provided (3 each from Donors 1 and 2; 33 and 41 donations from Donors 3 and 4, respectively).  All donors passed initial ARO screening and were well with no antimicrobial use nor travel history. Of the 80 donations, all were negative for MRSA, VRE, and CPO carriage but 3 (3.75%) were positive for ESBL Klebsiella pneumoniae [2 (6.1%) from Donor 3 and 1 (2.4%) from Donor 4]. The two positive results from Donor 3 were separated by 10 months. Conclusion:  Transient carriage of ESBLs in healthy pre-screened donors without illness, antimicrobial exposure, nor travel history suggests local transmission, possibly through food/water sources. Programs should consider screening all donations for ARO prior to acceptance into donor stool programs.