Use of bacteriophage in the treatment of experimental animal bacteremia from imipenem-resistant Pseudomonas aeruginosa.

The emergence of antibiotic-resistant bacterial strains still remains a significant problem for antimicrobial chemotherapy in the clinic. Bacterial viruses (bacteriophages or phages) have been suggested to be used as alternative therapeutic agents for bacterial infections. However, the efficacy of phage therapy in treating drug-resistant infections in humans is uncertain. Therefore in the present study, we examined the effectiveness of phages in the treatment of imipenem-resistant Pseudomonas aeruginosa (IMPR-Pa) infection in an experimental mouse model. Twenty-nine strains of phage were isolated from our hospital sewage, and phage ØA392 was representatively used for all testing because it has lytic activity against a wide range of clinical isolates of IMPR-Pa. We found that intraperitoneal (i.p.) injections of one IMPR-Pa strain (3 x 10(7) CFU) caused bacteremia and all mice died within 24 h. A single i.p. inoculation of the phage strain (MOI > or =0.01) at up to 60 min after the bacterial challenge was sufficient to rescue 100% of the animals. This lifesaving effect coincided with the rapid appearance of ØA392 in the circulation (within 2 h after i.p. injection), which remained at substantially higher levels for up to 48 h until the bacteria were eradicated. However, the survival rates of the mice dropped to approximately 50% and 20% when the same dose of this purified phage preparation was administered at 180 min and 360 min, respectively, after IMPR-Pa infections. In addition, we demonstrated that the ability of this phage to rescue bacteremic animals was due to the functional capabilities of the phage and not to a non-specific immune effect. The protection from death occurred only in animals inoculated with bacteria-specific virulent phage strains. When the heat-inactivated phages were used, the survival rate of the infected mice was dramatically reduced to 20% or lower. Moreover, the levels of the antibody against the phage were not significantly changed at the time when the bacteremic animals were protected by the active phages. Finally, our observations revealed that the inoculation of the mice with high-doses of ØA392 alone produced no adverse effects attributable to the phage. These data indicate that phages can save animals from pernicious P. aeruginosa infections and suggest that phage therapy may be potentially used as a stand-alone therapy for patients with IMPR-Pa infections.

Therapeutic effectiveness of bacteriophages in the rescue of mice with extended spectrum beta-lactamase-producing Escherichia coli bacteremia.

The emergence of multidrug-resistant bacteria has become a global crisis. Accumulating evidence shows that bacteriophages (phages) can rescue animals from a variety of lethal infections and be effective in treating drug-resistant infections in humans. Enterobacteriaceae, producing extended spectrum beta-lactamase enzymes (ESBLs), are resistant to a broad range of beta-lactamase antibiotics. One of the most common ESBL-producing gram-negative bacilli in Enterobacteriaceae is Escherichia coli. Since ESBL-producing E. coli poses a formidable challenge in the management of critically ill patients with bacterial infections, we undertook this study to explore the possible therapeutic utility of phages to control ESBL-producing E. coli infections. The phage Ø9882 used in this study was isolated from our hospital sewage and has lytic activity against a broad range of clinical isolates of ESBL-producing E. coli. ESBL-producing E. coli strains (n=30) were isolated in the clinic, and one of them was used to induce bacteremia in a murine model. Bacteremia was established by intraperitoneal (i.p.) injection of 3 x 10(7) CFU/ml, the minimum lethal dose (MLD) of bacterium in this animal model. Mice infected with the MLD of this strain alone died within 14 h, whereas a single i.p. inoculation of Ø9882 (MOI > or =10(-4)) given 40 min after the bacterial challenge led to 100% survival at 24-168 h, compared to 0% survival of saline-treated controls. Protection was obtained even when administration of the phage was delayed up to 60 min after the bacterial infection and the survival rate of infected animals was 60% at 168 h. Furthermore, it was shown that the therapeutic efficacy of Ø9882 in lethal systemic infection in our model is due to the functional capability of the phage and not the nonspecific immune effects. Our data both in vitro and in vivo revealed that: i) the protection of mice from death occurred only in animals infected with selected bacterial strains and the virulent phage specific to them; ii) when the phages were heat-inactivated, survival of the infected mice was strikingly decreased to 0; and iii) the level of antibody against the phage was not substantially elevated when the bacteremic animals were protected by the phage. The present findings indicate that phages can effectively rescue our mouse model from bacteremia and death, and thus provide the rationale and framework to evaluate the therapeutical efficacy of lytic phages against fatal ESBL-producing E. coli infections in humans.