Klebsiella Pneumoniae: Virulence, Biofilm and Antimicrobial Resistance

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Divya Bharathi

PIDJ ESPID Reports and Reviews PIDJ-217-384 CONTENTS Klebsiella pneumoniae Vitulence, Biofilm and Resistance in Klebsiella EDITORIAL BOARD Piperaki et al Editor: Delane Shingadia Board Members David Burgner (Melbourne, Cristiana Nascimento-Carvalho George Syrogiannopoulos XXX Australia) (Bahia, Brazil) (Larissa, Greece) Kow-Tong Chen (Tainan,Taiwan) Ville Peltola (Turku, Finland) Tobias Tenenbaum (Mannhein, Germany) Luisa Galli (Florence, Italy) Emmanuel Roilides (Thessaloniki, Marc Tebruegge (Southampton, UK) Steve Graham (Melbourne, Pediatr Infect Dis J Greece) Marceline Tutu van Furth (Amsterdam, Australia) Ira Shah (Mumbai, India) The Netherlands)

Lippincott Williams & Wilkins Klebsiella pneumoniae: Virulence, Biofilm and Antimicrobial

Hagerstown, MD Resistance

Evangelia-Theophano Piperaki, MD, PhD,* George A. Syrogiannopoulos, MD, PhD,† Leonidas S. Tzouvelekis, MD, PhD,* and George L. Daikos, MD, PhD‡

Key Words: Klebsiella pneumoniae, virulence, ingitis in premature neonates and infants as the immune response through cytokine and biofilm, resistance well as serious infections in immunocompro- chemokine production (Fig. 1). Among the mised and malnourished children, whereas effector cells that are recruited first to the in the community, K. pneumoniae is a com- infection site are the neutrophils. Impor- mon cause of urinary tract infections among tant mediators involved in this process are lebsiella pneumoniae is a ubiquitous immunocompetent children. interleukin (IL)-8 and IL-23, which induces K Gram-negative encapsulated bacterium In recent years, most K. pneumoniae production of IL-17 that promotes granu- that resides in the mucosal surfaces of mam- infections are caused by strains termed “clas- lopoietic response.6-7 IL-12 also amplifies the mals and the environment (soil, water, etc.). sic” K. pneumoniae (cKp). These strains expression of IL-17 through production of In humans, K. pneumoniae colonizes the persist in hospital environments and cause interferon-gamma. Other factors participat- gastrointestinal tract and less frequently the infections in debilitated patients. cKp strains ing in the immune response are the produc- 2017 nasopharynx, whence it gains entry to the cir- appear to be distinct from hypervirulent K. tion of IL-1β via activation of the NOD-like culation and other tissues causing infection. pneumoniae (hvKp), a variant that was first receptor pyrin domain containing (NLRP3) In the pre-antibiotic era, K. pneumoniae was described in the Asian Pacific Rim to cause inflammasome pathway and the production an important cause of community-acquired community-acquired, invasive and metastatic of other pro-inflammatory cytokines such as pneumonia, especially in alcoholics and dia- 8 Copyright © 2017 Wolters Kluwer Health, Inc. All rights infections, including liver abscess, endoph- tumor necrosis factor-a (TNF-α) and IL-6. betics. In the antibiotic era, K. pneumoniae thalmitis, meningitis and septic arthritis in The specific factors that K. pneumoniae reserved. became established in hospitals as a leading diabetics and immunocompetent young indi- employs to circumvent the immune defenses are cause of healthcare-associated infections.1 1002 2–5 not fully elucidated. Presently, there are 4 well- In pediatric wards, it causes sepsis and men- viduals. The emergence and spread of new multidrug-resistant (MDR) clones and the characterized virulence factors, namely the fim- international dissemination of hvKp strains briae, the capsule, the lipopolysaccharide (LPS) Accepted for publication May 9, 2017. and siderophores.6,9 K. pneumoniae is equipped 1005 From the *Department of Microbiology, School of have renewed interest in K. pneumoniae. Medicine, National and Kapodistrian University with adhesins, type 1 and type 3 fimbriae, which of Athens, Athens, Greece; †Department of Pedi- facilitate adherence to epithelial and immune 0891-3668 atrics, University of Thessaly, University General VIRULENCE cells as well as to abiotic surfaces (Fig. 1). Based Hospital of Larissa, Larissa, Greece; and ‡First To establish infection, K. pneumoniae Department of Medicine, National and Kapodis- on the composition of capsular polysaccharides trian University of Athens, Laikon General Hospi- must overcome mechanical and chemical (CPs) of K. pneumoniae, 78 distinct capsular 10.1097/INF.0000000000001675 tal, Athens, Greece. barriers and escape host humoral and cellular serotypes (K1 to K78) have been recognized. Of The authors have no funding or conflicts of interest innate defenses. After gaining access to the note, the vast majority of hvKp strains belong to disclose. host, the invading organisms are recognized Address for correspondence: George L. Daikos, MD, PhD, to serotypes K1 and K2. The high level of viru- The Pediatric Infectious Disease Journal First Department of Medicine, National and Kapodis- by the immune cells through the pattern lence of hvKp strains is, at least partially, due to trian University of Athens, Laikon General Hospital, recognition receptors and trigger produc- excess production of capsular material (hyper- Athens, Greece. E-mail: [email protected]. tion of various immune mediators. Central mucoviscous phenotype). Although the capsule Copyright © 2017 Wolters Kluwer Health, Inc. All 36 rights reserved. role in the innate immune response plays composition plays an important role in protect- ISSN: 0891-3668/17/3610-1002 the monocyte/macrophage system, which ing K. pneumoniae against the host immune DOI: 10.1097/INF.0000000000001675 has phagocytic capabilities and orchestrates response, it is likely that other factors contribute 10 TheThe ESPIDESPID Reports Reports and and Reviews Reviews of of Pediatric Pediatric Infectious Infectious Disease Diseases Journal series series topics, topics, authors authors and and contents contents are are chosen chosen and and approved approved independently by the Editorial Board of ESPID. October 1002 | www.pidj.com The Pediatric Infectious Disease Journal • Volume 36, Number 10, October 2017 2017 Copyright © 2017 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. The Pediatric Infectious Disease Journal • Volume 36, Number 10, October 2017 Vitulence, Biofilm and Resistance in Klebsiella

FIGURE 1. Schematic presentation of K. pneumoniae virulence factors and of host innate immune response. Capsular polysaccharides prevent phagocytosis and block complement-mediated lysis and opsonization. The intact LPS elicits a robust inflammatory response and prevents binding of C1q to bacteria and the subsequent activation of the complement pathway. Certain strains may modify the LPS making it unrecognizable to immune cells, whereas others may use the capsule to prevent LPS detection by toll-like receptor (TLR4) receptors. K. pneumoniae is equipped with types 1 and 3 fimbriae mediating adhesion to biotic and abiotic surfaces facilitating epithelial cell invasion and biofilm formation. It also synthesises siderophores (enterobactin, aerobactin, yersiniabactin and salmochelin) to acquire iron from the host. The monocyte/macrophage system plays a central role in the innate immune response, through phagocytosis and production of immune mediators such as cytokines and chemokines. Important mediator in this process is IL-23, that induces IL-17 production which along with IL-8 promote neutrophil recruit- ment. IL-17 expression is also amplified by IL-12 through IFN-γ. Other important cytokines are IL-1β, produced via activation of the NOD-like receptor pyrin domain-containing (NLRP3) inflammasome pathway, and other pro-inflammatory cytokines such as tumor necrosis factor-a (TNF-α) and IL-6. Perpendicular line (───│) indicates inhibition. Adapted from Paczosa and Mecsas.6 IFN-γ, interferon-gamma. to virulence as well. Indeed, certain K. pneumo- yersiniabactin are more prevalent in hvKp than are embedded within a self-produced matrix niae strains may modify the LPS to a degree cKp.6,9 Recent data suggest that hvKp strains of extracellular polymeric substance adhere that is not recognized by the host cells and oth- have the ability to produce larger amounts and to each other and/or to a surface.13 Extracel- ers may use capsule to mask LPS from detec- biologically more active iron-acquisition mole- lular polymeric substance is a complex struc- tion by toll-like receptor (TLR4) receptors.10,11 cules than nonvirulent strains, a mechanism that ture comprising polysaccharides, proteins These modifications dampen the inflammatory may contribute to virulence and pathogenesis.12 and DNA. The most clinically significant response and decrease bacterial clearance. Also, Although the clinical manifestations of infec- K. pneumoniae biofilms are those formed the ability of K. pneumoniae to steal iron from tions caused by hvKp strains have been well on the inner surfaces of catheters and other the host is critical for its growth and replication. described, as yet there is no single reliable bio- indwelling devices. K. pneumoniae biofilms Four iron-acquisition molecules (siderophores) logic marker that distinguishes these from other may also contribute to colonization of the have been described in K. pneumoniae: entero- K. pneumoniae strains. gastrointestinal, respiratory and urinary tract bactin, yersiniabactin, salmochelin and aerobac- and the development of invasive infections tin. Enterobactin has the highest affinity for iron, especially in immunocompromised patients. is present in both classical and hypervirulent BIOFILMS Development of K. pneumoniae bio- strains and is considered the primary iron uptake K. pneumoniae is able to form biofilms on solid surfaces proceeds from cell system. Unlike enterobactin, aerobactin and films, that is, aggregates in which cells that adherence, to formation of microcolonies,

maturation and finally dispersal of free-living colonization with multiple sequence types our treatment options are limited.25,26 Poly- cells. The most important surface structures (STs) of ESBL in the neonatal intensive care myxins (colistin and polymyxin B) are con- involved in the formation process are the unit, a recognized risk factor for develop- sidered among the most active agents against type 3 fimbriae and the CP.14 Fimbriae medi- ment of infection.16 In another study from the CP-Kp. To timely achieve the desired serum ate stable adherence, whereas CPs ultimately United States that analyzed resistance trends concentration, a loading dose of colistin is affect cell-to-cell communication and bio- in Enterobacteriaceae in children from 1999 required followed by adequate maintenance film architecture. Given the dynamic process to 2011, the prevalence of isolates exhibiting doses to attain optimal drug exposure.27 of biofilm production and the variability of resistance to third generation cephalosporins Aminoglycosides (gentamicin or amikacin) environmental stimuli, embedded cells must increased from 1.39% in 1999–2001 to 3% demonstrate good susceptibility profile to a be capable of swift and extensive changes in 2010–2011.17 Of note, although the K. proportion of CP-Kp, and this class of agents in gene expression. Transcriptional regula- pneumoniae isolates included in the analysis could serve as a backbone in the treatment of tion is controlled by quorum sensing, that were 7.7% of the total bacterial population CP-Kp infections, provided we exploit their is, a system of signals and responses that co- examined, they represented 33.1% of ESBL- well-studied pharmacokinetic/pharmacody- ordinate gene expression in a microorganism producing Enterobacteriaceae. Similarly namic (PK/PD) features and dose them opti- community. Putative quorum sensing–asso- with adult patients, in children, bloodstream mally. Treatment with tigecycline is another ciated regulators and autoinducers have been infections (BSIs) caused by ESBL-producing option, but standard dosing regimens of this described in K. pneumoniae,14 but available Enterobacteriaceae are associated with pro- agent attain suboptimal exposure to the drug; data are still incomplete. longed hospitalization and worse outcomes. this could be associated with adverse out- K. pneumoniae cells within bio- The dubious clinical efficacy of comes when it is used in nonapproved indi- films are partially protected from immune penicillin-inhibitor combinations and the cations, particularly in patients with hospital defenses. The matrix blocks the access of fact that a meaningful proportion of such acquired pneumonia/ventilator associated antibodies and antibacterial peptides and isolates have developed resistance to fluori- pneumonia (HAP/VAP) or BSIs. Moreover, reduces the efficiency of complement and nated quinolones and other antibiotics ren- this agent may cause permanent teeth dis- phagocytosis. Existence of mechanisms that dered carbapenems the first-line treatment coloration and enamel hypoplasia in children actively skew immunity toward reduced against ESBL-K. pneumoniae infections. <8 years of age, and its use should be limited inflammatory responses and establishment of The extensive use of carbapenems had as a only in those children in whom no other treat- chronic infection is also possible.15 consequence the emergence and rapid dis- ment option is available.26 Given that fosfo- What biofilms are most notorious for semination of MDR, carbapenemase-pro- mycin displays good in vitro activity against is high-level resistance to antibiotics. The ducing K. pneumoniae (CP-Kp) strains. The most CP-Kp, this agent could be selected as most important factor determining resistance prevalent carbapenemase types in K. pneu- salvage therapy to treat critically ill patients is bacterial growth status. Within the “inner moniae are the Klebsiella pneumoniae car- with infections caused by these pathogens. core” of a biofilm, bacteria are adapted to bapenemases (KPCs) (KPC-2 to KPC-13), The potential of fosfomycin to rapidly select starvation and low oxygen resulting in growth the metallo-β-lactamases (Verona integron- resistant mutants during therapy is a matter arrest that, in turn, diminishes the efficiency encoded metallo-β-lactamase, VIM; imi- of concern and this agent should be adminis- of antibiotics targeting metabolically active penemase, IMP; and New Delhi metallo-β- tered always with another active compound. and dividing cells. lactamase, NDM types) and the oxacillinase Although it sounds paradoxical, a propor- (OXA)-type enzymes (mainly OXA-48). tion of CP-Kp, depending on the geographic ANTIMICROBIAL RESISTANCE The carbapenemase-encoding genes are usu- region and the type of carbapenemase, exhib- K. pneumoniae strains are naturally ally carried on MDR transmissible plasmids its low carbapenem minimal inhibitory con- resistant to ampicillin, carbenicillin and that confer resistance to multiple antibiot- centration (MICs) (1–8 mg/L), and in patients ticarcillin because of production of a chro- ics.18 Three clones currently dominate the infected with such isolates, a high-dose/ mosomal penicillinase, sulfhydryl variable pandemic spread of CP-Kps: ST258, ST147 prolonged-infusion regimen of meropenem (SHV-1). What K. pneumoniae is notori- and ST15. Studies from different countries could drive the PK/PD profile of the drug ous for, however, is its propensity to collect show that hospitalized populations, includ- to acceptable exposure. Finally, the newly resistance plasmids. During the 1980s, K. ing patients from neonatal intensive care approved agent, ceftazidime/avibactam dem- pneumoniae became the index species for units, are rapidly and persistently colonized onstrates good in vitro activity against CP-Kp plasmids-encoding extended-spectrum-β- with K. pneumoniae ST258, especially in isolates that produce KPC and OXA-48 lactamases (ESBLs) conferring resistance to setting where the infection control practices enzymes, but not metallo-β-lactamases such expanded-spectrum cephalosporins. Initially, are inadequate. The majority of reports on as NDM, VIM or IMP. Nevertheless, clinical those were Temoniera (TEM)- and SHV- the dissemination of CP-Kp among chil- data on efficacy of ceftazidime–avibactam on type ESBLs and coexisted on the plasmids dren have been published from South-East infections caused by CP-Kp are scarce. with elements encoding resistance to ami- Asia and Western Pacific Regions.19–21 Data Despite the limitations in current evi- noglycosides, tetracyclines and trimetho- from pediatric patients in Europe, South and dence to guide treatment strategies against prim-sulfamethoxazole. The 1990s marked North America generally reflect the distribu- CP-Kp infections, as no randomized clinical the emergence of a new ESBL family, the tion of carbapenemase-producing Entero- trials have been performed to this date with cefotaximase-M (CTX-M) group, which bacteriaceae in adults.22–24 In the pediatric this objective, combination therapy appears are currently the dominant ESBLs in K. population, CP-Kp affect mainly critically ill to hold the most promise. Of note, the posi- pneumoniae. These enzymes confer resist- children with hematologic/oncologic condi- tive impact of combination therapy on sur- ance to penicillins and expanded-spectrum tions and an immunosuppressive state.22 The vival is more apparent in critically ill patients cephalosporins but are ineffective against most frequent types of infections are bactere- with severe infections.28,29 In low-risk BSIs carbapenems. Notably, ESBL-producing mia followed by respiratory and urinary tract and in nonbacteremic intra-abdominal or uri- organisms have disseminated in the pediat- infections. nary tract infections, combination schemes ric population as well. A recent study from Virtually, all CP-Kp isolates exhibit do not seem to provide any therapeutic Italy found increasing rates of intestinal extensive drug-resistance phenotypes, and advantage over monotherapy.29 Carbapenems

Copyright © 2017 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. The Pediatric Infectious Disease Journal • Volume 36, Number 10, October 2017 Vitulence, Biofilm and Resistance in Klebsiella

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