Acinetobacter baumannii has emerged as an important nosocomial pathogen.  Hospitals have reported emergence of multidrug resistance / Extensively drug resistant Acinetobacter species from various geographic areas, and this organism has become endemic in some of them. Acinetobacter species have been implicated in a wide range of infections such as pneumonia, skin and soft-tissue infections, wound infections, urinary tract infections, meningitis, and bloodstream infections. In 2011, WHO declared “combat drug resistance: no action today, no cure tomorrow.” In this context, utility of bacteriophage as an anti-infection modality was tried in animal wound model and also appearance of anti-phage antibodies was evaluated.

Acinetobacter baumannii isolated from the clinical specimens was found to be susceptible to colistin and 92% were considered as MDR strains and 64% as XDR and 88% were carbapenem-resistant. Bacteriophage active against clinical strains of A. baumannii was isolated from hospital sewage sludge. The bacteriophage isolated against A. baumannii formed plaques against XDR strains of A. baumannii. The utility of bacteriophage specific for A. baumannii to resolve wound infection was demonstrated in excision diabetic rabbit wound model.

The rabbits were randomly divided into 5 groups of five animals each. The upper lateral right hind limb of the rabbit was shaved and cleaned with spirit and an area of about 225 mm² of the skin was excised with full thickness using a sterile scissor under ketamine anaesthesia.

Group I : Excision wound was non-infected (Control).

Group II : Excision wound was infected with 400 µl of a 10⁸ XDR AB at 0 hr and observed for 6 weeks to study the course of the infection and clinical conditions of the infected animals.

Group III: Excision wound was infected with 400 µl of a 10⁸ XDR AB at 0 hr and after 48 hr of infection, the wound was debrided and challenged with local spray of 400 µl of 3X10⁹ PFU/ml Acinetobacter phage.

Group IV: Excision wound was infected with 400 µl of a 10⁸ XDR AB at 0 hr and after 48 hr of infection, challenged with intramuscular injection of 400 µl of antibiotic colistin (4 mg/kg body weight/day).

Group V: Excision Wound was non infected and sprayed with phage 400 µl of 3X10⁹ PFU/ml Acinetobacter phage (Phage Control).

The animals were observed for 6 weeks. Sequential cultures were done on all animals by collecting pus samples for Gram stain, culture and phage estimation as per the Clinical Microbiology Proficiency-Testing. A significant decrease in infection, period of epithelization, and wound contraction was observed in the phage-challenged group when compared with antibiotic-treated group and the control group. The antibody response was tested by phage neutralization test by collecting blood every week till 6 week followed by plaque assay. The anti-phage neutralizing antibodies was not observed even after 6^th week after administration of phages locally.

In this study, we were able to demonstrate phage prophylaxis against experimental XDR A. baumannii infections in uncontrolled diabetic rabbits similar to those that are common in humans. There was no evidence of general sepsis, and the rabbits, including phage controls, appeared remarkably well and also neutralizing antibodies against phages were not demonstrated indicating host immune system did not elicit immune response against phages used. The potential of phage therapy has been the subject of several recent scientific reviews, and this study reinforces the view that phage as a therapeutic agent is worth exploring when there are limited options for treatment as in a case of XDR Acinetobacter infection.

Key Words

XDR, Acinetobacter baumannii, Bacteriophage, Excision wound model