Next-Generation Sequencing and Comparative Analysis of Sequential Outbreaks Caused by Multidrug-Resistant Acinetobacter baumannii at a Large Academic Burn Center

Abstract

f Next-generation sequencing (NGS) analysis has emerged as a promising molecular epidemiological method for investigating health care-associated outbreaks. Here, we used NGS to investigate a 3-year outbreak of multidrug-resistant Acinetobacter baumannii (MDRAB) at a large academic burn center. A reference genome from the index case was generated using de novo assembly of PacBio reads. Forty-six MDRAB isolates were analyzed by pulsed-field gel electrophoresis (PFGE) and sequenced using an Illumina platform. After mapping to the index case reference genome, four samples were excluded due to low coverage, leaving 42 samples for further analysis. Multilocus sequence types (MLST) and the presence of acquired resistance genes were also determined from the sequencing data. A transmission network was inferred from genomic and epidemiological data using a Bayesian framework. Based on single-nucleotide variant (SNV) differences, this MDRAB outbreak represented three sequential outbreaks caused by distinct clones. The first and second outbreaks were caused by sequence type 2 (ST2), while the third outbreak was caused by ST79. For the second outbreak, the MLST and PFGE results were discordant. However, NGS-based SNV typing detected a recombination event and consequently enabled a more accurate phylogenetic analysis. The distribution of resistance genes varied among the three outbreaks. The first-and second-outbreak strains possessed a bla OXA-23-like group, while the thirdoutbreak strains harbored a bla OXA-40-like group. NGS-based analysis demonstrated the superior resolution of outbreak transmission networks for MDRAB and provided insight into the mechanisms of strain diversification between sequential outbreaks through recombination.H ealth care-associated infections (HAI) are a substantial cause of morbidity and mortality in acute-care hospitals (1). Acinetobacter baumannii is an important opportunistic pathogen causing HAI (2) and has become one of the most common colonizing pathogens in burn patients (3, 4). A. baumannii may cause serious outbreaks despite the implementation of rigorous infection prevention interventions and control measures, which occur most commonly in intensive care units (5-8). Furthermore, the emergence of multidrug-resistant A. baumannii (MDRAB), especially carbapenem-resistant A. baumannii (CRAB), has become a global concern (5, 9). Patients infected by multidrug-resistant Acinetobacter strains are likely to have higher mortality rates and longer lengths of hospitalization than those infected by susceptible strains (10).Pulsed-field gel electrophoresis (PFGE) has been the gold standard approach for bacterial strain typing in hospital outbreak investigations, but several disadvantages have been described, including that it is a time-consuming, labor-intensive, and technically demanding assay and that is has limited reproducibility among laboratories; also, the interpretation of the relative relatedness of strains using this method may be discordant and subjective (11-13). For these reasons, next...

Timeline of health care–associated infections and pathogens after burn injuries

“…878 potential pathogens were recovered from these 774 patients; 298 (5%) had 1 potential pathogen isolated, 164 (3%) had 2 potential pathogens isolated, 97 (2%) had 3 potential pathogens isolated, and 215 (4%) had 4 or more potential pathogens isolated. Of note, two outbreaks with Acinetobacter baumannii occurred in the time periods 9/2007 to 4/2008 and 1/2009 to 9/2010 15 .…”

Genomic Analysis of Multidrug-Resistant Escherichia coli from North Carolina Community Hospitals: Ongoing Circulation of CTX-M-Producing ST131- H 30Rx and ST131- H 30R1 Strains

“…Whole-genome sequencing (WGS), an emerging tool for advanced molecular epidemiological investigations involving MDR organisms, allows identification of extended resistance and virulence genotypes and elucidation of transmission routes (7)(8)(9)(10)(11)(12)(13). Recently, WGS-based phylogenetic analyses of E. coli ST131 strains identified a CTX-M-15-associated sublineage, designated H30Rx, within ST131-H30R, which is the fluoroquinoloneresistant subset within ST131-H30 (11).…”

The whole genome sequencing of Acinetobacter-calcoaceticus-baumannii complex strains involved in suspected outbreak in an Intensive Care Unit of a pediatric hospital

“…It is important to stress that these mutations have never been reported to induce polymyxins resistance in accordance to the phenotypic susceptibility tests (MIC ≤ 1 mg/ml). [18,19] Sulfonamide resistance in gram-negative bacteria is dependent on genes encoding dihydropteroate-synthase sul1 and sul2. [32] In our analysis we found that both ST195 isolates are characterized by the sul2 sulphonamide resistance determinant, while the ST281 strain presented the sul1 gene.…”

“…[16] To trace HAI it is necessary to use the best high resolution typing approach that unequivocally identifies Acinetobacter species and their clonality, in order to search for the index case and to define the strains relatedness to avoid nosocomial cross transmission. [17][18][19] This study described the use of whole genome sequencing (WGS) by Ion Torrent Personal Genome Machine to analyze Acinetobacter-calcoaceticusbaumannii complex strains isolated in a possible outbreak occurring in a Intensive Care Unit of a pediatric hospital. De novo assembled sequences from 13 strains were analyzed to define strains clonality and Resistoma.…”