Elsevier

Burns

Volume 43, Issue 7, November 2017, Pages 1532-1543
Burns

Liposome loaded phage cocktail: Enhanced therapeutic potential in resolving Klebsiella pneumoniae mediated burn wound infections

https://doi.org/10.1016/j.burns.2017.03.029Get rights and content

Highlights

  • Five phages sharing broad host range were used for preparation of a phage cocktail.

  • The phage cocktail was encapsulated in liposomes.

  • Liposomes increased phage retention time in vivo thus potentiating their efficacy.

  • Liposome entrapped phage cocktail was highly effective in reducing bacterial burden.

Abstract

Background

Klebsiella pneumoniae is one of the predominant pathogens in burn wound infections, and prevalence of multidrug resistant strains has further complicated the situation. An increased interest in phage therapy as a means of combating infection has been accruing in recent years. In order to overcome the drawbacks associated with phage therapy, the present study was conducted to evaluate the potential of liposomes as a delivery vehicle for phage in the treatment of burn wound infection.

Methods

Burn wound infection with Klebsiella pneumoniae B5055 was established in BALB/c mice. The therapeutic efficacy of free phage cocktail in comparison to liposome entrapped phage cocktail in resolving the course of burn wound infection in mice was evaluated.

Results

The results depicted that mice treated with liposomal entrapped phage cocktail showed higher reduction in bacterial load in blood and major organs. This was accompanied with faster resolution of the entire infection process as compared to non-liposomal free phage cocktail. The liposomes increased phage retention time in vivo thus potentiating efficacy. Liposomal phage preparation was able to protect all the test animals from death even when there was a delay of 24 h in instituting the therapy.

Conclusion

The results showed the potential of liposome entrapped phage cocktail for treating Klebsiella pneumoniae mediated infections. Thus, this strategy can serve as an effective approach for treating Klebsiella mediated burn wound infections in individuals who do not respond to conventional antibiotic therapy.

Introduction

Nosocomial infections due to Klebsiella pneumoniae are a major cause of morbidity and mortality among burn patients [1]. It is an important pathogen that accounts for 15.2% of all burn wound infections caused by gram-negative bacteria [2]. The propensity of this organism to disseminate promptly leaves little time to institute effective antimicrobial treatment. Moreover, with its ability to develop resistance to commonly employed antibiotics, management of K. pneumoniae especially in burn patients has become challenging [3]. Due to the inexorable spread of antibiotic resistant-bacteria and lack of development of new antibiotics, there is an urgent need to explore newer therapeutic options against this pathogen.

Phage therapy represents a safe and potent alternative strategy against drug resistant pathogens [4], [5]. Although success in the therapeutic use of bacteriophages for treating various bacterial infections has already been substantiated by many workers [6], [7], [8], [9] in the past but till date, none of the phage based therapies have successfully made it to the market. The intrinsic disadvantages associated with phage therapy need to be examined for its clinical success in near future. The major drawbacks include: high specificity, making them narrow range antibacterial agents [10], rapid clearance of phages by reticuloendothelial system of our body, poor pharmacokinetic profile that negatively affects the potency of treatment [11], [12]. Thirdly, most of the phage protection studies in animals have been performed with phages being administered immediately following bacterial infection, which is not the situation in a clinical setting [13].

In the present study, two major issues associated with phage therapy i.e. narrow host range and rapid clearance have been addressed by employing (i) phage cocktail (rather than monophage therapy) and (ii) suitable lipid based phage delivery system i.e. liposomes for enhancing the viability and stability of bacteriophages, both in vitro as well as in vivo. Among the plethora of delivery systems, liposomes are one of the most intensively employed system due to their nontoxic nature, biocompatibility with phages, non-immunogenic effect and GRAS status (generally regarded as safe) [14]. The present study for the first time provides information on the application of liposome encapsulated lytic phage cocktail for treating burn wound infection.

Section snippets

Bacterial strains

K. pneumoniae B5055 was used in the present study. It was procured from Dr. Matthias Trautmann, Department of Medical Microbiology and Hygiene, Ulm, Germany. Clinical isolates of K. pneumoniae were procured from Post Graduate Institute of Medical Education and Research (PGIMER) and Government Medical College and Hospital (GMCH-32) Chandigarh, India. Morphological and biochemical properties of the isolates were identified according to Bergey’s Manual of Systematic Bacteriology [15]. The strains

Preparation of liposomes

The liposomal vesicles encapsulating phage cocktail were prepared and DLS measurements of their size and polydispersity index performed. The results for size distribution indicated that liposomal formulation having PC: CHOL: T-80: SA in a ratio of 8:2:1:0.5 showed mean diameter of 229.83 nm for a PI value of 0.259. Phage cocktail encapsulation efficiency in the lipid vesicles was high (79.2% ± 5.6), probably because of positively charged surfactant stearyl amine that led to increase in the

Discussion

Therapeutic application of bacteriophages to control bacterial infections has rekindled interest in phage therapy as an alternative treatment option for eradicating drug resistant bacterial pathogens [6], [7], [9]. However, it still suffers from issues of poor stability, narrow spectrum and poor pharmacokinetic profile. There is a need to look into the use of Drug Delivery Systems (DDS) for stable and efficient delivery of lytic self-propagating bacterial viruses so as to circumvent associated

Conflict of interest

The authors report no conflict of interest.

Acknowledgement

The financial grant received from the Indian Council of Medical Research (ICMR), New Delhi, India, is gratefully acknowledged.

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