scholarly journals Phages from Ganges River curtail in vitro biofilms and planktonic growth of drug resistant Klebsiella pneumoniae in a zebrafish infection model

2021 ◽  
Author(s):  
Niranjana Sri Sundaramoor ◽  
Subramanian Thothathri ◽  
Muthu Meenakshi Bhaskaran ◽  
ArunKumar GaneshPrasad ◽  
Saisubramanian Nagarajan
Keyword(s):  
Klebsiella Pneumoniae ◽  
Infection Model ◽  
Histopathological Analysis ◽  
Drug Resistant ◽  
Ganges River ◽  
Promising Alternative ◽  
Planktonic Growth ◽  
Time Kill

Abstract Bacteriophages are a promising alternative for curtailing infections caused by multi drug resistant (MDR) bacteria. The objective of the present study is to evaluate phage populations from water bodies to inhibit planktonic and biofilm mode of growth of drug resistant Klebsiella pneumoniae in vitro and curtail planktonic growth in vivo in a zebrafish model. Phage specific to K. pneumoniae (MTCC 432) was isolated from Ganges River (designated as KpG). One-step growth curve, in vitro time kill curve study and in vivo infection model were performed to evaluate the ability of phage to curtail planktonic growth. Crystal violet assay and colony biofilm assay were performed to determine the action of phages on biofilms. KpG phages had a greater burst size, better bactericidal potential and enhanced inhibitory effect against biofilms formed at liquid air and solid air interfaces. In vitro time kill assay showed a 3 log decline and a 6 log decline in K. pneumoniae colony counts, when phages were administered individually and in combination with streptomycin, respectively. In vivo injection of KpG phages revealed that it did not pose any toxicity to zebrafish as evidenced by liver/brain enzyme profiles and by histopathological analysis. The muscle tissue of zebrafish, infected with K. pneumoniae and treated with KpG phages alone and in combination with streptomycin showed a significant 77.7% and 97.2 % decline in CFU/ml, respectively, relative to untreated control. Our study reveals that KpG phages has the potential to curtail plantonic and biofilm mode of growth in vivo in higher animal models.

scholarly journals Phages from Ganges River curtail in vitro biofilms and planktonic growth of drug resistant Klebsiella pneumoniae in a zebrafish infection model

2020 ◽  
Author(s):  
Niranjana Sri Sundaramoor ◽  
Subramanian Thothathri ◽  
Muthu Meenakshi Bhaskaran ◽  
ArunKumar GaneshPrasad ◽  
Saisubramanian Nagarajan
Keyword(s):  
Klebsiella Pneumoniae ◽  
Infection Model ◽  
Histopathological Analysis ◽  
Drug Resistant ◽  
Ganges River ◽  
Promising Alternative ◽  
Planktonic Growth ◽  
Time Kill

Abstract Bacteriophages are a promising alternative for curtailing infections caused by multi drug resistant (MDR) bacteria. The objective of the present study is to evaluate phage populations from water bodies to inhibit planktonic and biofilm mode of growth of drug resistant Klebsiella pneumoniae in vitro and curtail planktonic growth in vivo in a zebrafish model. Phage specific to K. pneumoniae (MTCC 432) was isolated from Ganges river (designated as KpG). One-step growth curve, in vitro time kill curve study and in vivo infection model were performed to evaluate the efficacy of phage to curtail planktonic growth. Crystal violet assay and colony biofilm assay was done to determine the action of phages on biofilms. KpG phages had a greater burst size, better bactericidal potential and enhanced inhibitory effect against biofilms formed at liquid air and solid air interfaces. In vivo injection of KpG phages revealed that it did not pose any toxicity to zebrafish as evidenced by liver/brain enzyme profiles and by histopathological analysis. In vitro time kill assay showed a 3 log decline and a 6 log decline in K. pneumoniae colony counts, when phages were administered individually and in combination with streptomycin, respectively. The muscle tissue of zebrafish, infected with K. pneumoniae and treated with KpG phages showed a significant 2 log decline in bacterial counts relative to untreated control. Our study reveals that KpG phages has the potential to curtail plantonic and biofilm mode of growth in vivo in higher animal models.

scholarly journals Phages from Ganges River curtail in vitro biofilms and planktonic growth of drug resistant Klebsiella pneumoniae in a zebrafish infection model

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Niranjana Sri Sundaramoorthy ◽  
Subramaniam Thothathri ◽  
Muthumeenakshi Bhaskaran ◽  
ArunKumar GaneshPrasad ◽  
Saisubramanian Nagarajan
Keyword(s):  
Klebsiella Pneumoniae ◽  
Infection Model ◽  
Histopathological Analysis ◽  
Drug Resistant ◽  
Ganges River ◽  
Promising Alternative ◽  
Planktonic Growth ◽  
Time Kill

AbstractBacteriophages are a promising alternative for curtailing infections caused by multi drug resistant (MDR) bacteria. The objective of the present study is to evaluate phage populations from water bodies to inhibit planktonic and biofilm mode of growth of drug resistant Klebsiella pneumoniae in vitro and curtail planktonic growth in vivo in a zebrafish model. Phage specific to K. pneumoniae (MTCC 432) was isolated from Ganges River (designated as KpG). One-step growth curve, in vitro time kill curve study and in vivo infection model were performed to evaluate the ability of phage to curtail planktonic growth. Crystal violet assay and colony biofilm assay were performed to determine the action of phages on biofilms. KpG phages had a greater burst size, better bactericidal potential and enhanced inhibitory effect against biofilms formed at liquid air and solid air interfaces. In vitro time kill assay showed a 3 log decline and a 6 log decline in K. pneumoniae colony counts, when phages were administered individually and in combination with streptomycin, respectively. In vivo injection of KpG phages revealed that it did not pose any toxicity to zebrafish as evidenced by liver/brain enzyme profiles and by histopathological analysis. The muscle tissue of zebrafish, infected with K. pneumoniae and treated with KpG phages alone and in combination with streptomycin showed a significant 77.7% and 97.2% decline in CFU/ml, respectively, relative to untreated control. Our study reveals that KpG phages has the potential to curtail plantonic and biofilm mode of growth in higher animal models.

scholarly journals Phages from Ganges River curtail in vitro biofilms and planktonic growth of drug resistant Klebsiella pneumoniae in a zebrafish infection model

2020 ◽  
Author(s):  
Niranjana Sri Sundaramoor ◽  
Subramanian Thothathri ◽  
Muthu Meenakshi Bhaskaran ◽  
ArunKumar GaneshPrasad ◽  
Saisubramanian Nagarajan
Keyword(s):  
Klebsiella Pneumoniae ◽  
Infection Model ◽  
Histopathological Analysis ◽  
Drug Resistant ◽  
Ganges River ◽  
Promising Alternative ◽  
Planktonic Growth ◽  
Time Kill

Abstract Bacteriophages are a promising alternative for curtailing infections caused by multi drug resistant (MDR) bacteria. The objective of the present study is to evaluate phage populations from water bodies to inhibit planktonic and biofilm mode of growth of drug resistant Klebsiella pneumoniae in vitro and curtail planktonic growth in vivo in a zebrafish model. Phage specific to K. pneumoniae (MTCC 432) was isolated from Ganges river (designated as KpG). One-step growth curve, in vitro time kill curve study and in vivo infection model were performed to evaluate the efficacy of phage against planktonic growth. Crystal violet assay and colony biofilm assay was done to determine the action of phages on biofilms. KpG phages had a greater burst size, better bactericidal potential and enhanced inhibitory effect against biofilms formed at liquid air and solid air interfaces. In vivo injection of KpG phages revealed that it did not pose any toxicity to zebrafish as evidenced by liver/brain enzyme profiles and by histopathological analysis. In vitro time kill assay showed a 3 log decline and a 6 log decline in K. pneumoniae colony counts, when phages were administered individually and in combination with streptomycin, respectively. The muscle tissue of zebrafish, infected with K. pneumoniae and treated with KpG phages showed a significant 2 log decline in bacterial counts relative to untreated control. Our study reveals that KpG phages has the potential to curtail plantonic and biofilm mode of growth in vivo in higher animal models.

Emergence of ceftazidime/avibactam resistance in KPC-3-producing Klebsiella pneumoniae in vivo

2019 ◽  
Vol 74 (11) ◽  
pp. 3211-3216 ◽  
Author(s):  
Stephan Göttig ◽  
Denia Frank ◽  
Eleonora Mungo ◽  
Anika Nolte ◽  
Michael Hogardt ◽  
...  
Keyword(s):  
Combination Therapy ◽  
Klebsiella Pneumoniae ◽  
Selection Pressure ◽  
Galleria Mellonella ◽  
Phenotypic Characterization ◽  
Infection Model ◽  
Development Of Resistance ◽  
Time Kill

Abstract Objectives The β-lactam/β-lactamase inhibitor combination ceftazidime/avibactam is active against KPC-producing Enterobacterales. Herein, we present molecular and phenotypic characterization of ceftazidime/avibactam resistance in KPC-3-producing Klebsiella pneumoniae that emerged in vivo and in vitro. Methods Sequence analysis of blaKPC-3 was performed from clinical and in vitro-generated ceftazidime/avibactam-resistant K. pneumoniae isolates. Time–kill kinetics and the Galleria mellonella infection model were applied to evaluate the activity of ceftazidime/avibactam and imipenem alone and in combination. Results The ceftazidime/avibactam-resistant clinical K. pneumoniae isolate revealed the amino acid change D179Y in KPC-3. Sixteen novel mutational changes in KPC-3 among in vitro-selected ceftazidime/avibactam-resistant isolates were described. Time–kill kinetics showed the emergence of a resistant subpopulation under selection pressure with either imipenem or ceftazidime/avibactam. However, combined selection pressure with imipenem plus ceftazidime/avibactam prevented the development of resistance and resulted in bactericidal activity. Concordantly, the G. mellonella infection model revealed that monotherapy with ceftazidime/avibactam is prone to select for resistance in vivo and that combination therapy with imipenem results in significantly better survival. Conclusions Ceftazidime/avibactam is a valuable antibiotic against MDR and carbapenem-resistant Enterobacterales. Based on time–kill kinetics as well as an in vivo infection model we postulate a combination therapy of ceftazidime/avibactam and imipenem as a strategy to prevent the development of ceftazidime/avibactam resistance in KPC-producing Enterobacterales in vivo.

scholarly journals Polymyxin Triple Combinations against Polymyxin-Resistant, Multidrug-Resistant, KPC-Producing Klebsiella pneumoniae

2020 ◽  
Vol 64 (8) ◽  
Author(s):  
Su Mon Aye ◽  
Irene Galani ◽  
Heidi Yu ◽  
Jiping Wang ◽  
Ke Chen ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Triple Therapy ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Infection Model ◽  
Bacterial Killing ◽  
Standard Dosage ◽  
Time Kill

ABSTRACT Resistance to polymyxin antibiotics is increasing. Without new antibiotic classes, combination therapy is often required. We systematically investigated bacterial killing with polymyxin-based combinations against multidrug-resistant (including polymyxin-resistant), carbapenemase-producing Klebsiella pneumoniae. Monotherapies and double- and triple-combination therapies were compared to identify the most efficacious treatment using static time-kill studies (24 h, six isolates), an in vitro pharmacokinetic/pharmacodynamic model (IVM; 48 h, two isolates), and the mouse thigh infection model (24 h, six isolates). In static time-kill studies, all monotherapies (polymyxin B, rifampin, amikacin, meropenem, or minocycline) were ineffective. Initial bacterial killing was enhanced with various polymyxin B-containing double combinations; however, substantial regrowth occurred in most cases by 24 h. Most polymyxin B-containing triple combinations provided greater and more sustained killing than double combinations. Standard dosage regimens of polymyxin B (2.5 mg/kg of body weight/day), rifampin (600 mg every 12 h), and amikacin (7.5 mg/kg every 12 h) were simulated in the IVM. Against isolate ATH 16, no viable bacteria were detected across 5 to 25 h with triple therapy, with regrowth to ∼2-log10 CFU/ml occurring at 48 h. Against isolate BD 32, rapid initial killing of ∼3.5-log10 CFU/ml at 5 h was followed by a slow decline to ∼2-log10 CFU/ml at 48 h. In infected mice, polymyxin B monotherapy (60 mg/kg/day) generally was ineffective. With triple therapy (polymyxin B at 60 mg/kg/day, rifampin at 120 mg/kg/day, and amikacin at 300 mg/kg/day), at 24 h there was an ∼1.7-log10 CFU/thigh reduction compared to the starting inoculum for all six isolates. Our results demonstrate that the polymyxin B-rifampin-amikacin combination significantly enhanced in vitro and in vivo bacterial killing, providing important information for the optimization of polymyxin-based combinations in patients.

scholarly journals In VitroandIn VivoActivities of OP0595, a New Diazabicyclooctane, against CTX-M-15-Positive Escherichia coli and KPC-Positive Klebsiella pneumoniae

2016 ◽  
Vol 60 (5) ◽  
pp. 3001-3006 ◽  
Author(s):  
Akihiro Morinaka ◽  
Yuko Tsutsumi ◽  
Keiko Yamada ◽  
Yoshihiro Takayama ◽  
Shiro Sakakibara ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Pneumoniae ◽  
Infection Model ◽  
Gram Negative Bacteria ◽  
Content Type ◽  
Effective Combination ◽  
Time Kill ◽  
Lactamase Inhibitor

ABSTRACTGram-negative bacteria are evolving to produce β-lactamases of increasing diversity that challenge antimicrobial chemotherapy. OP0595 is a new diazabicyclooctane serine β-lactamase inhibitor which acts also as an antibiotic and as a β-lactamase-independent β-lactam “enhancer” againstEnterobacteriaceae. Here we determined the optimal concentration of OP0595 in combination with piperacillin, cefepime, and meropenem, in addition to the antibacterial activity of OP0595 alone and in combination with cefepime, inin vitrotime-kill studies and anin vivoinfection model against five strains of CTX-M-15-positiveEscherichia coliand five strains of KPC-positiveKlebsiella pneumoniae. An OP0595 concentration of 4 μg/ml was found to be sufficient for an effective combination with all three β-lactam agents. In bothin vitrotime-kill studies and anin vivomodel of infection, cefepime-OP0595 showed stronger efficacy than cefepime alone against all β-lactamase-positive strains tested, whereas OP0595 alone showed weaker or no efficacy. Taken together, these data indicate that combinational use of OP0595 and a β-lactam agent is important to exert the antimicrobial functions of OP0595.

scholarly journals XDR-Klebsiella pneumoniae isolates harboring blaOXA-48: In vitro and in vivo evaluation using a murine thigh-infection model

2019 ◽  
Vol 244 (18) ◽  
pp. 1658-1664
Author(s):  
Noha A Kamel ◽  
Wafaa N El-Tayeb ◽  
Mona R El-Ansary ◽  
Mohamed T Mansour ◽  
Khaled M Aboshanab
Keyword(s):  
Klebsiella Pneumoniae ◽  
Synergistic Effect ◽  
Cancer Patients ◽  
Bacterial Count ◽  
Infection Model ◽  
Drug Resistant ◽  
Extensively Drug Resistant ◽  
Pediatric Cancer Patients

Blood stream infection with extensively drug-resistant-carbapenamase producing Klebsiella (K.) pneumoniae usually represents a major threat with medical challenges among hospitalized cancer patients with poor functional status and underlying diseases. Accordingly, the aim of the study was to evaluate the efficacy of different antibiotics either alone or in combinations against extensively drug-resistant-OXA-48 producing K. pneumoniae clinical isolates that were previously recovered from febrile neutropenic pediatric cancer patients. The antimicrobial activity of amikacin, gentamicin, colistin, ertapenem, imipenem, meropenem and tigecycline was assessed by broth microdilution method. The results revealed that all the tested OXA-48 producing K. pneumoniae isolates exhibited extensively drug-resistant phenotype and all of them were susceptible to tigecycline. Checkerboard method was used to determine the fraction inhibitory concentration index, to further classify the effect of antibiotic combination as synergistic, additive, indifferent, or antagonistic effect. The results revealed that in vitro dual carbapenem combination of ertapenem with meropenem had shown synergistic effect against all of the tested isolates. Additionally, synergistic effect of meropenem with colistin was detected among three of four isolates tested. Herein we investigated the in vivo activity of colistin, meropenem alone and in combination in a rat thigh infection model. The results showed that addition of meropenem to colistin was not effective at reduction of bacterial count as compared to colistin alone at 24 h post treatment. Accordingly, we can conclude that in vitro antibiotic combinations of dual carbapenems (ertapenem plus meropenem) and meropenem plus colistin showed synergism in 100% and 75% of the tested isolates, respectively. Colistin alone had significantly reduced bacterial count while its combination with meropenem was not superior to monotherapy in murine thigh infection model. Impact statement The present study aimed to evaluate the effectiveness of various antibiotics both in vitro and in vivo using murine animal model either alone or in combination against various strains of extensively drug-resistant (XDR) Klebsiella pneumoniae, life-threatening pathogens of relevant medical importance isolated from febrile neutropenic pediatric cancer patients. This work also emphasizes how to select the appropriate antibiotics options and help the physicians to choice the appropriate antibiotic for the treatment of such superbugs (extensively drug-resistant (XDR) Klebsiella pneumoniae). The results showed that in vitro dual carbapenem combination of ertapenem with meropenem had shown synergistic effect against all of the tested XDR isolates. Antibiotic combinations of dual carbapenems and meropenem plus colistin showed synergism in 100% and 75% of the testes isolates, respectively. Results of the in vivo evaluation, colistin alone had significantly reduced bacterial count while its combination with meropenem was not superior to monotherapy.

scholarly journals Bactericidal Activities of Meropenem and Ertapenem against Extended-Spectrum-β-Lactamase-Producing Escherichia coli and Klebsiella pneumoniae in a Neutropenic Mouse Thigh Model

2007 ◽  
Vol 51 (4) ◽  
pp. 1481-1486 ◽  
Author(s):  
C. Andrew DeRyke ◽  
Mary Anne Banevicius ◽  
Hong Wei Fan ◽  
David P. Nicolau
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Pneumoniae ◽  
Infection Model ◽  
Bacterial Reduction ◽  
Efficacy Analysis ◽  
Percent Time ◽  
Extended Spectrum ◽  
Wide Range

ABSTRACT The purpose of this study was to examine the in vivo efficacies of meropenem and ertapenem against extended-spectrum-β-lactamase (ESBL)-producing isolates with a wide range of MICs. Human-simulated dosing regimens in mice were designed to approximate the free drug percent time above the MIC (fT>MIC) observed for humans following meropenem at 1 g every 8 h and ertapenem at 1 g every 24 h. An in vivo neutropenic mouse thigh infection model was used to examine the bactericidal effects against 31 clinical ESBL Escherichia coli and Klebsiella pneumoniae isolates and 2 non-ESBL isolates included for comparison at a standard 105 inoculum. Three isolates were examined at a high 107 inoculum as well. Meropenem displayed greater in vitro potency, with a median MIC (range) (μg/ml) of 0.125 (0.03 to 32), than did ertapenem, with 0.5 (0.012 to 128). Seven of the 31 ESBL isolates were removed from the efficacy analysis due to their inability to establish infection in the mouse model. When MICs were ≤1.5 μg/ml for ertapenem (≤0.5 μg/ml for meropenem), similar reductions in CFU (≈ 2-log kill) were observed for both ertapenem (fT>MIC ≥ 23%) and meropenem (fT>MIC ≥ 75%). Ertapenem showed bacterial regrowth for seven of eight isolates, with MICs of ≥2 μg/ml (fT>MIC ≤ 20%), while meropenem displayed antibacterial potency that varied from a static effect to a 1-log bacterial reduction in these isolates (fT>MIC = 30 to 65%). At a 107 inoculum, both agents eradicated bacteria due to adequate exposures (fT>MIC = 20 to 45%). Due to low MICs, no difference in bacterial kill was noted for the majority of ESBL isolates tested. However, for isolates with raised ertapenem MICs of ≥2 μg/ml, meropenem displayed sustained efficacy due to its greater in vitro potency and higher resultant fT>MIC.

scholarly journals Klebsiella virus UPM2146 lyses multiple drug-resistant Klebsiella pneumoniae in vitro and in vivo

PLoS ONE ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. e0245354
Author(s):  
Omar Assafiri ◽  
Adelene Ai-Lian Song ◽  
Geok Hun Tan ◽  
Irwan Hanish ◽  
Amalia Mohd Hashim ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Multiple Drug ◽  
Drug Resistant ◽  
Zebrafish Model ◽  
Whole Genome Analysis ◽  
Narrow Host Range ◽  
Opportunistic Bacteria ◽  
Multiple Drug Resistant

Klebsiella pneumoniae are opportunistic bacteria found in the gut. In recent years they have been associated with nosocomial infections. The increased incidence of multiple drug-resistant K. pneumoniae makes it necessary to find new alternatives to treat the disease. In this study, phage UPM2146 was isolated from a polluted lake which can lyse its host K. pneumoniae ATCC BAA-2146. Observation from TEM shows that UPM2146 belongs to Caudoviriales (Order) based on morphological appearance. Whole genome analysis of UPM2146 showed that its genome comprises 160,795 bp encoding for 214 putative open reading frames (ORFs). Phylogenetic analysis revealed that the phage belongs to Ackermannviridae (Family) under the Caudoviriales. UPM2146 produces clear plaques with high titers of 1010 PFU/ml. The phage has an adsorption period of 4 min, latent period of 20 min, rise period of 5 min, and releases approximately 20 PFU/ bacteria at Multiplicity of Infection (MOI) of 0.001. UPM2146 has a narrow host-range and can lyse 5 out of 22 K. pneumoniae isolates (22.72%) based on spot test and efficiency of plating (EOP). The zebrafish larvae model was used to test the efficacy of UPM2146 in lysing its host. Based on colony forming unit counts, UPM2146 was able to completely lyse its host at 10 hours onwards. Moreover, we show that the phage is safe to be used in the treatment against K. pneumoniae infections in the zebrafish model.

scholarly journals Tridecaptin M, a New Variant Discovered in Mud Bacterium, Shows Activity against Colistin- and Extremely Drug-ResistantEnterobacteriaceae

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
Manoj Jangra ◽  
Manpreet Kaur ◽  
Rushikesh Tambat ◽  
Rohit Rana ◽  
Sushil K. Maurya ◽  
...  
Keyword(s):  
Hemolytic Activity ◽  
World Health ◽  
Fourth Generation ◽  
Infection Model ◽  
Drug Resistant ◽  
Content Type ◽  
Extensively Drug Resistant ◽  
New Variant

ABSTRACTThe World Health Organization has categorized the Gram-negative superbugs, which are inherently impervious to many antibiotics, as critical priority pathogens due to the lack of effective treatments. The breach in our last-resort antibiotic (i.e., colistin) by extensively drug-resistant and pan-drug-resistantEnterobacteriaceaestrains demands the immediate development of new therapies. In the present study, we report the discovery of tridecaptin M, a new addition to the family, and its potential against colistin-resistantEnterobacteriaceae in vitroandin vivo. Also, we performed mode-of-action studies using various fluorescent probes and studied the hemolytic activity and mammalian cytotoxicity in two cell lines. Tridecaptin M displayed strong antibacterial activity (MICs of 2 to 8 μg ml−1) against clinical strains ofKlebsiella pneumoniae(which were resistant to colistin, carbapenems, third- and fourth-generation cephalosporins, fluoroquinolones, fosfomycin, and other antibiotics) andmcr-1-positiveEscherichia colistrains. Unlike polymyxins, tridecaptin M did not permeabilize the outer membrane or cytoplasmic membrane. It blocked ATP synthesis in bacteria by dissipating the proton motive force. The compound exhibited negligible acquired resistance, lowin vitrocytotoxicity and hemolytic activity, and no significant acute toxicity in mice. It also showed promising efficacy in a thigh infection model of colistin-resistantK. pneumoniae. Altogether, these results demonstrate the future prospects of this class of antibiotics to address the unmet medical need to circumvent colistin resistance in extensively drug-resistantEnterobacteriaceaeinfections. The work also emphasizes the importance of natural products in our shrunken drug discovery pipeline.

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