Bacterial infections are normally treated with antibiotic chemotherapy. A growing number of infections, however, are becoming more difficult to treat as the antibiotics commonly prescribed are becoming less effective due to antibiotic resistance. Biofilm formation represents another strategy that allows bacteria to survive antibiotic treatment. Moreover, bacteria such as Staphylococcus aureus can polymerize host fibrinogen into a mesh of fibrin, forming a physical barrier to host immune cells. Strategies that disrupt the fibrin protecting the bacteria could support the resolution of the infection. In general, infections associated with medical implants can be extremely difficult to eradicate and can negatively affect patient's health. To counteract the bacterial threat, alternative approaches need to be considered and examined.
In this study we tested the activity of a staphylokinase phage-derived enzyme with / without support of antibiotic treatment on S. aureus cells. Staphylokinase is an enzyme that converts human plasminogen into plasmin with ultimate action of digesting fibrin. Bacteria were grown between 2 layers of collagen gel in presence of human plasma for a total of 16 hrs to form fully developed microcolonies surrounded by fibrin. After microscopic confirmation of the formation of the fibrin protective layer, staphylokinase (10 - 5 - 2 - 0.5µM) was applied to the cells for 120 min, followed by 4 hours gentamicin or vancomycin treatment (10xMIC). To determine the activity of each treatment, microscope images were taken to assess the fibrin degradation followed by quantification of bacteria count after antibiotic administration.
Microscopy evaluation showed that the fibrin protective structure was degraded by the activity of the staphylokinase with the strongest effect observed at the highest concentration. The activity of the enzyme alone resulted in a slightly reduction of the CFU count when compared to the untreated sample. A combination of 10 µg/ml gentamicin with 10 µM staphylokinase showed a 2-log fold reduction on the CFU count indicating a strong synergism of the paired antimicrobials, compared with gentamicin alone which did not affect the viability of the cells. Highest concentration of staphylokinase associated with 20 µg/ml vancomycin presented a moderate reduction of the CFU count, however not as efficient as observed for the gentamicin.
Our data suggest the importance of the staphylokinase treatment in disrupting the protective fibrine layer for maximizing the antibiotic action. Combination of specific enzymes and antibiotics could improve the resolution of difficult infections and prevent bacteria from developing antimicrobial resistance.