Lab Matters Summer 2018 - Page 42

APHL 2018 Annual Meeting Poster Abstracts Reliable Identification Methods for Carbapenem-resistant Carbapenemase-producing Acinetobacter (CP–CRA) Laboratory Surveillance of Enterobacteriaceae Isolated from Patients in Tennessee J. Dale and P. Snippes-Vagnone, Minnesota Department of Health Public Health Laboratory, St. Paul, MN (complete abstract in Infectious Disease, p. 76) Antibiotic resistance is a worldwide concern impacting the healthcare system with difficult to treat infections. An emerging threat is resistance to the carbapenem class of antibiotics, which has resulted in surveillance efforts to understand the prevalence and epidemiology of carbapenem resistant organisms. Acinetobacter baumannii is a hospital-associated, opportunistic pathogen with both intrinsic and acquired antibiotic resistance mechanisms. Carbapenemase-producing carbapenem-resistant Acinetobacter spp. (CP-CRA) often harbor carbapenemase genes on a plasmid that has the potential to transfer between bacterial species and genera. The prevalence of CP-CRA containing plasmid- encoded carbapenemases is poorly understood and the clinical significance is unclear. Therefore, it is imperative that efforts are taken to detect the carbapenem resistance mechanisms of CP-CRA to aid in infection control measures. Fundamental to treatment and control of CP-CRA are phenotypic and nucleic acid-based methods for carbapenemase detection. Here we used established phenotypic methods for carbapenemase detection, in conjunction with real- time PCR (RT-PCR) for Acinetobacter blaOXA ß-lactamase genes, to identify correlations between phenotype and genotype and provide insight into the clinical significance of CP-CRA. The carbapenem inactivation method (CIM) and modified CIM (mCIM) are phenotypic tests performed routinely for the detection of carbapenemase production in Enterobacteriaceae; however, these methods are unreliable for use with Acinetobacter spp. Therefore, we opted to re- examine the interpretive guidelines for carbapenemase production of CRA by comparing CIM and mCIM results with data obtained from RT-PCR detecting the presence of Acinetobacter plasmid- associated blaOXA genes (OXA-23, -24/40, -58) and the intrinsic blaOXA gene (OXA-51). We concur with the literature that the mCIM is not reliable for detecting CP-CRA. However, our data demonstrates that the interpretive criteria for CIM positive Acinetobacter spp. should include any zone size with colonies spread throughout the zone of inhibition. Use of the new interpretation strongly correlates w ith the identification of an Acinetobacter blaOXA gene. There are some isolates that harbor a blaOXA gene, but are CIM negative, requiring further molecular testing to determine gene expression levels that could explain the negative phenotype. In addition, the chromosomal or plasmid location of each blaOXA gene needs to be determined along with its capability for horizontal gene transfer (HGT). Overall, our data provide strong evidence that CIM results are a reliable indicator for CP-CRA and accurately predict the presence of a blaOXA gene. The identification of a CP-CRA isolate harboring a blaOXA gene, which may be plasmid-associated, is highly important considering the possibility of HGT to other pathogens. Genomic Investigation of a Protracted Carbapenem- Resistant Enterobacter aerogenes Outbreak in a Cardiac ICU at a Tertiary Care Center in Rochester, New York Presenter: Jennifer Dale, PhD, Minnesota Department of Health Public Health Laboratory, St. Paul, MN, Phone: 612.201.5043, Email: Comparison of ETEST ® and Broth Microdilution Methods for Antimicrobial Susceptibility Testing of Shigella sp. Isolates in New York City (complete abstract in Food Safety, p. 58) 40 LAB MATTERS Summer 2018 A. Malek, S. Taffner, H. Mostafa, J. Wang, S. Petry, L. Fine, P. Graman, D. Hardy, N. Pecora; University of Rochester Medical Center, Rochester, NY Background: Enterobacter spp. are significant nosocomial pathogens associated with outbreaks in intensive care units (ICUs). Between Jun-Oct 2017, carbapenem-resistant Enterobacter aerogenes (CR-EA) strains were isolated from patients in our cardiac ICU (CICU). Whole genome sequencing (WGS) of CR-EA isolates was undertaken to investigate patient-to-patient transmission, assess phylogeny relative to global strains and characterize molecular determinants of resistance and virulence. Methods: 22 CR-EA strains (12 outbreak, 10 other wards) were sequenced (Illumina Miseq) and investigated for phylogenetic relatedness by whole genome multi-locus sequence typing (wgMLST) using Ridom ® Seqsphere+ and the CFSAN SNP pipeline. To establish phylogeny with global E. aerogenes strains, 113 publically-available sequences were used for comparison using HARVEST genomics suite. Markers for antibiotic resistance and virulence factors were identified using curated databases. Results: WgMLST and core-SNP analyses revealed every CICU CR- EA isolate to be part of a single clonal cluster, grouping distantly from strains isolated from other wards and previous years. Barring a single 2015 strain, harboring an nmcAR locus, none of the CR- EA strains isolated from our hospital harbored genes encoding carbapenemases. Some CICU strains harbored mutations resulting in premature stop codons in outer membrane porin genes (omp36), likely contributing to the carbapenem resistant phenotype. Virulome analysis revealed the CICU strains to harbor genes encoding yersiniabactin and colibactin systems on the ICEKp10 pathogenicity island. Conclusions: Carbapenem-resistant Enterobacteriaceae are a major public health concern, with rapid spread attributed to the production of mobilizable carbapenemases. Here, we describe a hospital ward outbreak involving a clonal group of E. aerogenes carbapenemase non-producing strains. Virulence and resistance determinants underlying infections by Enterobacter spp. are poorly understood and the latter may involve mechanisms such as porin disruption and/or AmpC cephalosporinase overproduction, which present complex diagnostic and management challenges. Yersiniabactin and colibactin systems have been implicated in the invasiveness of hyper-virulent Klebsiella pneumoniae. Intact clusters of these components in the CICU outbreak strains may indicate a role for these loci in enhanced survival and effective transmission in a subset of E. aerogenes lineages. Prospective WGS presents a powerful resource to complement traditional epidemiology in helping hospitals and public health institutions track transmission events and assess the effectiveness of control measures in real-time. Presenter: Adel Malek, University of Rochester Medical Center, Department of Pathology and Laboratory Medicine, Rochester, NY, Email: PublicHealthLabs @APHL