Carbapenem-Resistant Klebsiella pneumoniae Exhibit Variability in Capsular Polysaccharide and Capsule Associated Virulence Traits

Journal of Infectious Diseases, Sep 2014

Background. Novel therapies are urgently needed to treat carbapenem-resistant Klebsiella pneumoniae (CR-Kp)-mediated infection, which constitute a major health threat in the United States. In order to assess if it is feasible to develop anticapsular antibodies as a potential novel therapy, it is crucial to first systematically characterize capsular polysaccharide (CPS) and virulence traits in these strains. Methods. Forty CR-Kp were genotyped by pulsed field gel electrophoresis, multilocus sequence typing (MLST), and molecular capsule typing (C-patterns and wzi sequencing). Their biofilm formation, serum resistance, macrophage-mediated killing, and virulence in Galleria mellonella were compared. MAb (1C9) was generated by co-immunization with 2 CPSs, and cross-reactivity was investigated. Results. MLST assigned 80% of CR-Kp isolates to the ST258-clone. Molecular capsule typing identified new C-patterns, including C200/wzi-154, which was widely represented and associated with blaKPC-3-bearing strains. Heterogeneity was detected in biofilm formation and macrophage-mediated killing. Differences in serum resistance correlated with virulence in G. mellonella. ST258 strains carrying blaKPC-3 were less virulent than those with blaKPC-2. MAb 1C9 cross-reacted with 58% of CR-Kp CPSs. Conclusions. CR-Kp ST258 strains exhibit variability of virulence-associated traits. Differences were associated with the type of KPC gene and CPS. Identification of cross-reacting anti-CPS mAbs encourages their development as adjunctive therapy.

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Carbapenem-Resistant Klebsiella pneumoniae Exhibit Variability in Capsular Polysaccharide and Capsule Associated Virulence Traits

JID Carbapenem-Resistant Klebsiella pneumoniae Exhibit Variability in Capsular Polysaccharide and Capsule Associated Virulence Traits Elizabeth Diago-Navarro 2 Liang Chen 1 Virginie Passet 0 4 Seth Burack 2 Amaia Ulacia-Hernando 2 Rosy Priya Kodiyanplakkal 2 Michael H. Levi 3 Sylvain Brisse 0 4 Barry N. Kreiswirth 1 Bettina C. Fries () 2 0 Institut Pasteur , Microbial Evolutionary Genomics 1 Public Health Research Institute Tuberculosis Center, NJMS-Rutgers University , Newark, New Jersey 2 Department of Medicine Infectious Disease Division Albert Einstein College of Medicine and Montefiore Medical Center , Bronx , New York 3 Department of Clinical Microbiology Montefiore Medical Center , Bronx , New York 4 CNRS, UMR3525 , Paris , France Background. Novel therapies are urgently needed to treat carbapenem-resistant Klebsiella pneumoniae (CRKp)-mediated infection, which constitute a major health threat in the United States. In order to assess if it is feasible to develop anticapsular antibodies as a potential novel therapy, it is crucial to first systematically characterize capsular polysaccharide (CPS) and virulence traits in these strains. Methods. Forty CR-Kp were genotyped by pulsed field gel electrophoresis, multilocus sequence typing (MLST), and molecular capsule typing (C-patterns and wzi sequencing). Their biofilm formation, serum resistance, macrophage-mediated killing, and virulence in Galleria mellonella were compared. MAb (1C9) was generated by coimmunization with 2 CPSs, and cross-reactivity was investigated. Results. MLST assigned 80% of CR-Kp isolates to the ST258-clone. Molecular capsule typing identified new C-patterns, including C200/wzi-154, which was widely represented and associated with blaKPC-3-bearing strains. Heterogeneity was detected in biofilm formation and macrophage-mediated killing. Differences in serum resistance correlated with virulence in G. mellonella. ST258 strains carrying blaKPC-3 were less virulent than those with blaKPC-2. MAb 1C9 cross-reacted with 58% of CR-Kp CPSs. Conclusions. CR-Kp ST258 strains exhibit variability of virulence-associated traits. Differences were associated with the type of KPC gene and CPS. Identification of cross-reacting anti-CPS mAbs encourages their development as adjunctive therapy. - In the past decades carbapenem-resistant Klebsiella pneumoniae (CR-Kp) strains have emerged in the United States since 2001 and worldwide [1]. Currently, the most common carbapenemase in the United States is K. pneumoniae carbapenemase (KPC), an Ambler molecular class A enzyme that facilitates hydrolysis of a broad variety of β-lactams. Recent CDC surveillance data estimate that the prevalence of CR-Kp in the United States among healthcare-associated infections increased from 1.6% in 2001 to 10.4% in 2011 [2]. The majority of clinical CR-Kp isolates in the United States are of MLST–defined clonal background ST258 that carry KPCs (blaKPC-2 or blaKPC-3) [3]. CR-Kp infections have high mortality rates (40%–50%) and result in increased treatment and hospitalization costs [4, 5]. With no novel antimicrobials for emerging CR-Kp in sight, efforts to explore alternative treatment options and prevention of global dissemination are warranted [6]. One of the main virulence factors of K. pneumoniae is its capsular polysaccharide (CPS) [7]. CPS is expressed in vivo, promotes biofilm formation, and exerts an anti-opsonic effect, all of which evade the host immune response. Strategies targeting the CPS have been successful both in vaccine development as well as passive immunotherapy for other encapsulated pathogens. For K. pneumoniae protective efficacy of anticapsular antibodies has been demonstrated in animal models, further supporting efforts to develop antibodies as adjunctive therapy [8]. CPS genes in K. pneumoniae strains are chromosomally encoded and clustered in the cps genomic locus [9, 10]. Over 77 capsular (K) serotypes have been described. However, strains of ST258 have not been extensively characterized for their Kserotype or molecular methods of cps cluster analysis such as C-pattern [10] and wzi sequencing [11]. In this study we characterized 40 CR-Kp strains from the Bronx with respect to their CPS, biofilm formation, resistance to serum and macrophage killing, as well as virulence in a Galleria mellonella and mouse model. This study is the first to our knowledge to document significant CPS-associated variability including novel C-patterns and wzi alleles among CR-Kp strains of the ST258 clone. Despite variability, cross-reactive antibodies could be generated. In addition, significant variability was documented with respect to virulence-associated traits. The implications of these findings for efforts of developing anti-capsular antibodies are discussed. MATERIAL AND METHODS K. pneumoniae Strains CR-Kp strains were collected from inpatients at Montefiore Medical Center (MMC) in Bronx, New York, that presented with CR-Kp bacteremia between December 2010 (...truncated)


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Elizabeth Diago-Navarro, Liang Chen, Virginie Passet, Seth Burack, Amaia Ulacia-Hernando, Rosy Priya Kodiyanplakkal, Michael H. Levi, Sylvain Brisse, Barry N. Kreiswirth, Bettina C. Fries. Carbapenem-Resistant Klebsiella pneumoniae Exhibit Variability in Capsular Polysaccharide and Capsule Associated Virulence Traits, Journal of Infectious Diseases, 2014, pp. 803-813, 210/5, DOI: 10.1093/infdis/jiu157