Graves, James, Nathan Yaldo, Cody Bruno, Stefan Petrovski, Elizabeth Stahl, and Monir Mardini

Pseudomonas aeruginosa is a highly antibiotic resistant opportunistic pathogen. Phage (bacterial viruses) could be considered as an antimicrobial agent for use in some types of infections. However, mechanisms may exist for bacteria to avoid killing by viruses just as has occurred with the antibiotics. In this work, P. aeruginosa survivors of phage in a biofilm assay were examined. The host strain P. aeruginosa ATCC 13388 designated suitable for biodegradation studies and medical research was able to grow up to the edge of 3 out of 5 different antibiotic discs used in an evaluation of sensitivity. The phage was isolated from the sediment of a tributary of the Detroit River. In the routine test dilution (RTD), to determine the highest titer of phage to give complete lysis, performed on brain heart infusion (BHI) agar, as the multiplicity of infection (MOI) decreased, opacity of the lytic zones increased even though the sites were infected by a high number of phage. The lowest number of phage to give complete lysis was about 10 raised to the power of 4 plaque forming units (PFU). Reducing the cell population density did not appear to have a considerable effect in the RTD assay. The Gram stain of bacteria from lytic zones of apparent complete lysis revealed the presence of gram-negative rods. Colonies of bacteria produced from samples from lytic zones by streak plate culture were similar in appearance to those for the original strain. In a cytochrome oxidase test colonies demonstrated the dark purple pigment characteristic of a positive reaction. The Enterotube II, which is a multiple biochemical test system made of 12 test chambers in a series, showed that the survivors were nonfermentative. Survivor bacteria did not appear to produce phage. Survivors could be missed target cells, phase variants, mutants, revertants or lysogens (cells that maintain the virus internally). It is Important to consider the potential significance of survivors of bacteria after phage therapy.