scholarly journals 1952. Bacteriophage Treatment Improves Survival of Mice Infected with Carbapenem-Resistant Klebsiella pneumoniae

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S60-S60
Author(s):  
Shayla Hesse ◽  
Natalia Malachowa ◽  
Adeline Porter ◽  
Brett Freedman ◽  
Scott Kobayashi ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Bacterial Infections ◽  
Phage Therapy ◽  
Control Group ◽  
Lytic Phage ◽  
Mouse Blood ◽  
Carbapenem Resistant ◽  
In Vitro Data

Abstract Background Bacteriophage (phage) therapy is being considered as a treatment option for patients with multi-drug-resistant bacterial infections. However, there is a dearth of controlled clinical data to support therapeutic phage efficacy. As a first step toward addressing this deficiency, we tested the ability of two well-characterized phages, alone and in combination, to kill carbapenem-resistant Klebsiella pneumoniae (ST258) in blood in vitro and rescue mice from lethal ST258 infection. Methods Wild-type C57BL/6J mice were infected with a lethal inoculum of ST258 by intra-peritoneal (IP) injection followed 1 hour later by IP administration of lytic phage P1, P2, or P1+P2 at a multiplicity of infection (MOI) estimated at 1. Survival of each group of mice was tracked for 10 days. In separate experiments, mice were sacrificed at 1 hour, 24 hours, and 48 hours post-phage treatment. Mouse blood and tissues were collected at each timepoint for enumeration of bacteria and phage, screening for phage resistance, and histopathology. Results ST258 survival in mouse blood in vitro was significantly less after 1 hour of incubation with P1 or P1+P2 (MOI 1) compared with the control group (no phage). Consistent with the in vitro data, none of the mice (0/15) in the control group (no phage) survived to 10 days post-infection, whereas 12/15, 14/15, and 15/15 mice survived in the P2, P1, and P1+P2-treated groups, respectively (P < 0.0001). Conclusion Prompt, systemic administration of lytic bacteriophages rescued mice from lethal ST258 infection. These data support the potential of phage therapy to effectively treat infections caused by ST258. It will be important to assess whether, for other phage-bacteria combinations, in vitro lysis in blood correlates with in vivo treatment efficacy and therefore may have predictive utility. Disclosures All Authors: No reported Disclosures.

scholarly journals Topically Applied Bacteriophage to Control Multi-Drug Resistant Klebsiella pneumoniae Infected Wound in a Rat Model

Antibiotics ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 1048
Author(s):  
Mohamed S. Fayez ◽  
Toka A. Hakim ◽  
Mona M. Agwa ◽  
Mohamed Abdelmoteleb ◽  
Rania G. Aly ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Rat Model ◽  
Bacterial Infections ◽  
Control Group ◽  
Lytic Phage ◽  
Drug Resistant ◽  
Infected Wound ◽  
Specific Phage

(Background): Multi-drug-resistant Klebsiella pneumoniae (MDR-KP) has steadily grown beyond antibiotic control. Wound infection kills many patients each year, due to the entry of multi-drug resistant (MDR) bacterial pathogens into the skin gaps. However, a bacteriophage (phage) is considered to be a potential antibiotic alternative for treating bacterial infections. This research aims at isolating and characterizing a specific phage and evaluate its topical activity against MDR-KP isolated from infected wounds. (Methods): A lytic phage ZCKP8 was isolated by using a clinical isolate KP/15 as a host strain then characterized. Additionally, phage was assessed for its in vitro host range, temperature, ultraviolet (UV), and pH sensitivity. The therapeutic efficiency of phage suspension and a phage-impeded gel vehicle were assessed in vivo against a K. pneumoniae infected wound on a rat model. (Result): The phage produced a clear plaque and was classified as Siphoviridae. The phage inhibited KP/15 growth in vitro in a dose-dependent pattern and it was found to resist high temperature (˂70 °C) and was primarily active at pH 5; moreover, it showed UV stability for 45 min. Phage-treated K. pneumoniae inoculated wounds showed the highest healing efficiency by lowering the infection. The quality of the regenerated skin was evidenced via histological examination compared to the untreated control group. (Conclusions): This research represents the evidence of effective phage therapy against MDR-KP.

scholarly journals Safety and Efficacy of a Phage, kpssk3, in an in vivo Model of Carbapenem-Resistant Hypermucoviscous Klebsiella pneumoniae Bacteremia

2021 ◽  
Vol 12 ◽  
Author(s):  
Yunlong Shi ◽  
Yuan Peng ◽  
Yixin Zhang ◽  
Yu Chen ◽  
Cheng Zhang ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Gut Microbiota ◽  
Mammalian Cells ◽  
Phage Therapy ◽  
In Vivo Model ◽  
Therapeutic Effects ◽  
Global Public Health ◽  
Carbapenem Resistant

Antimicrobial resistance (AMR) is one of the most significant threats to global public health. As antibiotic failure is increasing, phages are gradually becoming important agents in the post-antibiotic era. In this study, the therapeutic effects and safety of kpssk3, a previously isolated phage infecting carbapenem-resistant hypermucoviscous Klebsiella pneumoniae (CR-HMKP), were evaluated in a mouse model of systemic CR-HMKP infection. The therapeutic efficacy experiment showed that intraperitoneal injection with a single dose of phage kpssk3 (1 × 107 PFU/mouse) 3 h post infection protected 100% of BALB/c mice against bacteremia induced by intraperitoneal challenge with a 2 × LD100 dose of NY03, a CR-HMKP clinical isolate. In addition, mice were treated with antibiotics from three classes (polymyxin B, tigecycline, and ceftazidime/avibactam plus aztreonam), and the 7 days survival rates of the treated mice were 20, 20, and 90%, respectively. The safety test consisted of 2 parts: determining the cytotoxicity of kpssk3 and evaluating the short- and long-term impacts of phage therapy on the mouse gut microbiota. Phage kpssk3 was shown to not be cytotoxic to mammalian cells in vitro or in vivo. Fecal samples were collected from the phage-treated mice at 3 time points before (0 day) and after (3 and 10 days) phage therapy to study the change in the gut microbiome via high-throughput 16S rDNA sequence analysis, which revealed no notable alterations in the gut microbiota except for decreases in the Chao1 and ACE indexes.

scholarly journals A Frameshift Mutation in wcaJ Associated with Phage Resistance in Klebsiella pneumoniae

2020 ◽  
Vol 8 (3) ◽  
pp. 378 ◽  
Author(s):  
Demeng Tan ◽  
Yiyuan Zhang ◽  
Jinhong Qin ◽  
Shuai Le ◽  
Jingmin Gu ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Phage Therapy ◽  
Rapid Development ◽  
Phage Resistance ◽  
Comparative Genomic ◽  
Lytic Phage ◽  
Wild Type ◽  
Coding Region

Phage therapy is a potential and promising avenue for controlling the emergence and spread of multidrug-resistant (MDR) Klebsiella pneumoniae, however, the rapid development of anti-phage resistance has been identified as an obstacle to the development of phage therapy. Little is known about the mechanism employed by MDR K. pneumoniae strains and how they protect themselves from lytic phage predation in vitro and in vivo. In this study, comparative genomic analysis shows undecaprenyl-phosphate glucose-1-phosphate transferase (WcaJ), the initial enzyme catalyzing the biosynthesis of colanic acid, is necessary for the adsorption of phage 117 (Podoviridae) to the host strain Kp36 to complete its lytic life cycle. In-frame deletion of wcaJ alone was sufficient to provide phage 117 resistance in the Kp36 wild-type strain. Complementation assays demonstrated the susceptibility of phage 117, and the mucoid phenotype could be restored in the resistant strain Kp36-117R by expressing the wild-type version of wcaJ. Remarkably, we found that bacterial mobile genetic elements (insA and insB) block phage 117 infections by disrupting the coding region of wcaJ, thus preventing phage adsorption to its phage receptor. Further, we revealed that the wcaJ mutation likely occurred spontaneously rather than adapted by phage 117 predation under unfavorable environments. Taken together, our results address a crucial evolutionary question around the mechanisms of phage–host interactions, increasing our current understandings of anti-phage defense mechanisms in this important MDR pathogen.

scholarly journals Comparative analysis of Clinial Carbapenem- Resistant (CR), Hypermucoviscous (HM) and CR-HM Klebsiella pneumoniae isolates in China

2020 ◽  
Author(s):  
Jun-Ying Zhu ◽  
Guang-Yu Wang ◽  
Qing Wei ◽  
Zhen Shen ◽  
Qiong Li ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Virulence Genes ◽  
Carbapenem Resistance ◽  
Resistance Rate ◽  
Molecular Characteristics ◽  
Carbapenem Resistant ◽  
Recent Emergence ◽  
String Test

Abstract Background: Although carbapenem-resistant Klebsiella pneumoniae (CRKP) and hypermucoviscous K. pneumoniae (HMKP) were largely non-overlapping, the recent emergence of CR-HMKP has raised great alarm in the world. We compared the molecular characteristics of CRKP, HMKP and CR-HMKP isolates.Results: 220 cases of K. pneumoniae isolates was collected and identified between Jan 2015 and Dec 2016 from Renji Hospital. Carbapenem resistance test and string test were performed to screen CRKP, HMKP and CR-HMKP isolates. All the CRKP, HMKP and CR-HMKP isolates were investigated for capsular genotyping, virulence genes and resistance genes by PCR and DNA sequencing. Multilocus sequence typing (MLST) was used to characterize isolates sequence types (STs). Serum killing assay and mouse lethality assay were respectively performed to confirm the virulence of the isolates in vitro and in vivo. Of 220 K. pneumoniae，71 HMKP, 84 CRKP and 8 CR-HMKP were identified. Resistance rate to carbapenems was significantly higher in CRKP than HMKP and CR-HMKP. For MLST and serotyping, ST23 (26.8%)，K1 (33.8%) and K2 (23.9%) serotypes were the most common in HMKP isolates while ST11 (84.5%, 100%) and K-nontypable (91.6%, 100%) were the predominant types in CRKP and CR-HMKP isolates. The existence of virulence genes rmpA, magA and iutA was significantly higher in HMKP while the prevalence of resistance gene blaKPC-2 was higher in CRKP and CR-HMKP. Virulence test in vivo and in vitro both showed the lower virulence of CRKP and CR-HMKP compared to HMKP.Conclusions: In spite of low virulence, the emergence of CR-HMKP indicates a confluence of hypermucoviscous phenotype and carbapenem resistance. Furthermore, the similar molecular characteristics between CRKP and CR-HMKP suggested that CR-HMKP might evolve from CRKP.

scholarly journals Serum Antibody Responses against Carbapenem-Resistant Klebsiella pneumoniae in Infected Patients

mSphere ◽  
2021 ◽  
Vol 6 (2) ◽  
Author(s):  
Kasturi Banerjee ◽  
Michael P. Motley ◽  
Elizabeth Diago-Navarro ◽  
Bettina C. Fries
Keyword(s):  
Klebsiella Pneumoniae ◽  
Capsular Polysaccharide ◽  
Serum Antibody ◽  
Antibody Responses ◽  
Capsular Polysaccharides ◽  
Content Type ◽  
Carbapenem Resistant ◽  
Reactive Properties

ABSTRACT Capsular polysaccharide (CPS) heterogeneity within carbapenem-resistant Klebsiella pneumoniae (CR-Kp) strain sequence type 258 (ST258) must be considered when developing CPS-based vaccines. Here, we sought to characterize CPS-specific antibody responses elicited by CR-Kp-infected patients. Plasma and bacterial isolates were collected from 33 hospital patients with positive CR-Kp cultures. Isolate capsules were typed by wzi sequencing. Reactivity and measures of efficacy of patient antibodies were studied against 3 prevalent CR-Kp CPS types (wzi29, wzi154, and wzi50). High IgG titers against wzi154 and wzi50 CPS were documented in 79% of infected patients. Patient-derived (PD) IgGs agglutinated CR-Kp and limited growth better than naive IgG and promoted phagocytosis of strains across the serotype isolated from their donors. Additionally, poly-IgG from wzi50 and wzi154 patients promoted phagocytosis of nonconcordant CR-Kp serotypes. Such effects were lost when poly-IgG was depleted of CPS-specific IgG. Additionally, mice infected with wzi50, wzi154, and wzi29 CR-Kp strains preopsonized with wzi50 patient-derived IgG exhibited lower lung CFU than controls. Depletion of wzi50 antibodies (Abs) reversed this effect in wzi50 and wzi154 infections, whereas wzi154 Ab depletion reduced poly-IgG efficacy against wzi29 CR-Kp. We are the first to report cross-reactive properties of CPS-specific Abs from CR-Kp patients through both in vitro and in vivo models. IMPORTANCE Carbapenem-resistant Klebsiella pneumoniae is a rapidly emerging public health threat that can cause fatal infections in up to 50% of affected patients. Due to its resistance to nearly all antimicrobials, development of alternate therapies like antibodies and vaccines is urgently needed. Capsular polysaccharides constitute important targets, as they are crucial for Klebsiella pneumoniae pathogenesis. Capsular polysaccharides are very diverse and, therefore, studying the host’s capsule-type specific antibodies is crucial to develop effective anti-CPS immunotherapies. In this study, we are the first to characterize humoral responses in infected patients against carbapenem-resistant Klebsiella pneumoniae expressing different wzi capsule types. This study is the first to report the efficacy of cross-reactive properties of CPS-specific Abs in both in vitro and in vivo models.

In vitro and in vivo synergistic effects of tigecycline combined with aminoglycosides on carbapenem-resistant Klebsiella pneumoniae

2021 ◽  
Author(s):  
Wentao Ni ◽  
Deqing Yang ◽  
Jie Guan ◽  
Wen Xi ◽  
Dexun Zhou ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Synergistic Effects ◽  
Molecular Characteristics ◽  
Synergistic Activity ◽  
Significant Activity ◽  
Clinical Pharmacokinetics ◽  
Carbapenem Resistant ◽  
Time Kill

Abstract Objectives Carbapenem-resistant Klebsiella pneumoniae (CR-KP) infections represent severe threats to public health worldwide. The aim of this study was to assess potential synergistic interaction between tigecycline and aminoglycosides via in vitro and in vivo studies. Methods Antibiotic resistance profiles and molecular characteristics of 168 CR-KP clinical isolates were investigated by susceptibility testing, PCR and MLST. Chequerboard tests and time–kill assays were performed for 20 CR-KP isolates to evaluate in vitro synergistic effects of tigecycline combined with aminoglycosides. A tissue-cage infection model of rats was established to evaluate in vivo synergistic effects. Different doses of tigecycline and aminoglycosides alone or in combination were administered for 7 days via tail vein injection. Antibiotic efficacy was evaluated in tissue-cage fluid and emergence of resistance was screened. Results The chequerboard tests showed that this combination displayed synergistic or partial synergistic activity against CR-KP. The time–kill assays further demonstrated that strong synergistic effects of such a combination existed against isolates that were susceptible to both drugs but for resistant isolates no synergy was observed if clinical pharmacokinetics were taken into consideration. The in vivo study showed that the therapeutic effectiveness of combination therapies was better than that of monotherapy for susceptible isolates, suggesting in vivo synergistic effects. Furthermore, combinations of tigecycline with an aminoglycoside showed significant activity in reducing the occurrence of tigecycline-resistant mutants. Conclusions Compared with single drugs, tigecycline combined with aminoglycosides could exert synergistic effects and reduce the emergence of tigecycline resistance. Such a combination might be an effective alternative when treating CR-KP infections in clinical practice.

scholarly journals Combination of Colistin and Azidothymidine Demonstrates Synergistic Activity against Colistin-Resistant, Carbapenem-Resistant Klebsiella pneumoniae

2020 ◽  
Vol 8 (12) ◽  
pp. 1964
Author(s):  
Ya-Ting Chang ◽  
Tsung-Ying Yang ◽  
Po-Liang Lu ◽  
Shang-Yi Lin ◽  
Liang-Chun Wang ◽  
...  
Keyword(s):  
Combination Therapy ◽  
Klebsiella Pneumoniae ◽  
Therapeutic Option ◽  
Rapid Evolution ◽  
Toxicity Testing ◽  
World Health ◽  
Synergistic Activity ◽  
Carbapenem Resistant

Carbapenem-resistant Enterobacteriaceae (CRE) is listed as an urgent threat by the World Health Organization because of the limited therapeutic options, rapid evolution of resistance mechanisms, and worldwide dissemination. Colistin is a common backbone agent among the “last-resort” antibiotics for CRE; however, its emerging resistance among CRE has taken the present dilemma to the next level. Azidothymidine (AZT), a thymidine analog used to treat human immunodeficiency virus/acquired immunodeficiency syndrome, has been known to possess antibacterial effects against Enterobacteriaceae. In this study, we investigated the combined effects of AZT and colistin in 40 clinical isolates of colistin-resistant, carbapenem-resistant K. pneumoniae (CCRKP). Eleven of the 40 isolates harbored Klebsiella pneumoniae carbapenemase. The in vitro checkerboard method and in vivo nematode killing assay both revealed synergistic activity between the two agents, with fractional inhibitory concentration indexes of ≤0.5 in every strain. Additionally, a significantly lower hazard ratio was observed for the nematodes treated with combination therapy (0.288; p < 0.0001) compared with either AZT or colistin treatment. Toxicity testing indicated potentially low toxicity of the combination therapy. Thus, the AZT–colistin combination could be a potentially favorable therapeutic option for treating CCRKP.

scholarly journals Novel Lytic Phages Protect Cells and Mice against Pseudomonas aeruginosa Infection

2021 ◽  
Author(s):  
Feng Chen ◽  
Xingjun Cheng ◽  
Jianbo Li ◽  
Xiefang Yuan ◽  
Xiuhua Huang ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Mouse Models ◽  
Bacterial Infections ◽  
Phage Therapy ◽  
Inflammatory Responses ◽  
Multi Drug Resistance ◽  
Bacterial Strains ◽  
Genetic Traits

With the fast emergence of serious antibiotic resistance and the lagged discovery of novel antibacterial drugs, phage therapy for pathogenic bacterial infections has acquired great attention in the clinics. However, development of therapeutic phages also faces tough challenges, such as laborious screening and time to generate effective phage drugs since each phage may only lyse a narrow scope of bacterial strains. Identifying highly effective phages with broad host ranges is crucial for improving phage therapy. Here, we isolated and characterized several lytic phages from various environments specific for Pseudomonas aeruginosa by testing their growth, invasion, host ranges, and potential for killing targeted bacteria. Importantly, we identified several therapeutic phages (HX1, PPY9, and TH15) with broad host ranges to lyse laboratory strains and clinical isolates of P. aeruginosa with multi-drug resistance (MDR) both in vitro and in mouse models. In addition, we analyzed critical genetic traits related to the high-level broad host coverages by genome sequencing and subsequent computational analysis against known phages. Collectively, our findings establish that these novel phages may have potential for further development as therapeutic options for patients who fail to respond to conventional treatments. IMPORTANCE Novel lytic phages isolated from various environmental settings were systematically characterized for their critical genetic traits, morphology structures, host ranges against laboratory strains and clinical multi-drug resistant (MDR) Pseudomonas aeruginosa, and antibacterial capacity both in vitro and in mouse models. First, we characterized the genetic traits and compared with other existing phages. Furthermore, we utilized acute pneumonia induced by laboratorial strain PAO1, and W19, an MDR clinical isolate and chronic pneumonia by agar beads laden with FDR1, a mucoid phenotype strain isolated from the sputum of a cystic fibrosis (CF) patient. Consequently, we found that these phages not only suppress bacteria in vitro but also significantly reduce the infection symptom and disease progression in vivo, including lowered bug burdens, inflammatory responses and lung injury in mice, suggesting that they may be further developed as therapeutic agents against MDR P. aeruginosa.

scholarly journals Microcin MccI47 selectively inhibits enteric bacteria and reduces carbapenem-resistant Klebsiella pneumoniae colonization in vivo when administered via an engineered live biotherapeutic

2021 ◽  
Author(s):  
Benedikt M Mortzfeld ◽  
Jacob D Palmer ◽  
Shakti K Bhattarai ◽  
Haley L Dupre ◽  
Regino Mercado-Lubo ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Genetically Modify ◽  
Enteric Bacteria ◽  
Probiotic Bacterium ◽  
Multicopy Plasmid ◽  
Gi Tract ◽  
Carbapenem Resistant ◽  
Antimicrobial Resistance Genes

Background: The gastrointestinal (GI) tract is the reservoir for multidrug-resistant (MDR) pathogens, specifically carbapenem-resistant (CR) Klebsiella pneumoniae and other Enterobacteriaceae, which often lead to the spread of antimicrobial resistance genes, severe extraintestinal infections, and lethal outcomes. Selective GI decolonization has been proposed as a new strategy for preventing transmission to other body sites and minimizing spreading to susceptible individuals. Results: Here, we purify the to-date uncharacterized class IIb microcin I47 (MccI47) and demonstrate potent inhibition of numerous Enterobacteriaceae, including MDR clinical isolates, in vitro at concentrations resembling those of commonly prescribed antibiotics. We then genetically modify the probiotic bacterium Escherichia coli Nissle 1917 (EcN) to produce MccI47 from a stable multicopy plasmid by using MccI47 toxin production in a counterselection mechanism to engineer one of the native EcN plasmids, which renders provisions for inducible expression and plasmid selection unnecessary. We then test the clinical relevance of the MccI47-producing engineered EcN in a murine CR K. pneumoniae colonization model and demonstrate significant MccI47-dependent reduction of CR K. pneumoniae abundance after seven days of daily oral live biotherapeutic administration without disruption of the resident microbiota. Conclusions: This study provides the first demonstration of MccI47 as a potent antimicrobial against certain Enterobacteriaceae, and its ability to significantly reduce the abundance of CR K. pneumoniae in a preclinical animal model, when delivered from an engineered live biotherapeutic product. This study serves as the foundational step towards the use of engineered live biotherapeutic products aimed at the selective removal of MDR pathogens from the GI tract.

scholarly journals Curcumin-meropenem synergy in carbapenem resistant Klebsiella pneumoniae curcumin-meropenem synergy

2021 ◽  
Author(s):  
Dumrul Gülen ◽  
Birol Şafak ◽  
Berna Erdal ◽  
Betül Günaydın
Keyword(s):  
Klebsiella Pneumoniae ◽  
Bacterial Infections ◽  
Efflux Pump ◽  
Synergistic Effects ◽  
Treatment Options ◽  
Resistance Pattern ◽  
Carbapenem Resistant ◽  
Efflux Pump Inhibition ◽  
Disk Method

Background and Objectives: The frequency of multiple resistant bacterial infections, including carbapenems, is increasing worldwide. As the decrease in treatment options causes difficulties in treatment, interest in new antimicrobials is increasing. One of the promising natural ingredients is curcumin. It is known to be effective in bacteria such as Pseudomonas aeruginosa, Escherichia coli, Burkholderia pseudomallei through efflux pump inhibition, toxin inhibition and enzymes. However, because its bioavailability is poor, it seffectiveness occurs in combination with antibiotics. In the study, the interaction of meropenem and curcumin in carbapenemase producing strains of Klebsiella pneumoniae was tested. Materials and Methods: Thirty-nine Klebsiella pneumoniae isolates, resistant to meropenem, were used in this study. From those 15 MBL, 6 KPC, 17 OXA-48 and 1 AmpC resistance pattern were detected by combination disk method. Meropenem and Curcumin MIC values were determined by liquid microdilution. Checkerboard liquid microdilution was used to determine the synergy between meropenem and curcumin. Results: Synergistic effects were observed in 4 isolates producing MBL, 3 isolates producing KPC, 4 isolates producing OXA-48, and 1 isolates producing AmpC (totally 12 isolates) according to the calculated FICI. No antagonistic effects were observed in any isolates. Conclusion: Curcumin was thought to be an alternative antimicrobial in combination therapies that would positively contribute to the treatment of bacterial infection. The effectiveness of this combination should be confirmed by other in vitro and clinical studies.

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