1114 - Combined application of bacterial predation and violacein to kill polymicrobial pathogenic communities

Abstract

Violacein is a bisindole antibiotic that is effective against Gram-positive bacteria while the bacterial predator, Bdellovibrio bacteriovorus HD100, predates on Gram-negative strains. In this study, we evaluated the use of both together against multidrug resistant pathogens. The two antibacterial agents did not antagonize the activity of the other. For example, treatment of Staphylococcus aureus with violacein reduced its viability by more than 2,000-fold with or without B. bacteriovorus addition. Likewise, predation of Acinetobacter baumannii reduced the viability of this pathogen by more than 13,000-fold, regardless if violacein was present or not. When used individually against mixed bacterial cultures containing both Gram-positive and Gram-negative strains, violacein and B. bacteriovorus HD100 were effective against only their respective strains. The combined application of both violacein and B. bacteriovorus HD100, however, reduced the total pathogen numbers by as much as 84,500-fold. Their combined effectiveness was also demonstrated using a 4-species culture containing S. aureus, A. baumannii, Bacillus cereus and Klebsiella pneumoniae. When used alone, violacein and bacterial predation reduced the total population by only 19% and 68%, respectively. In conjunction with each other, the pathogen viability was reduced by 2,965-fold (99.98%), illustrating the prospective use of these two antimicrobials together against mixed species populations. Within nature, bacteria are found primarily within polymicrobial communities. This is also true of infections, which may include both pathogens and commensal microbes 1,2. One benefit that bacteria within these communities gain is an increased resistance to antibiotics 3,4. Two reasons for this include the presence of resistant strains 5,6 , which can degrade the antibiotic, or because the other strains act as a "sink" for the antibiotic, diluting its impact on the susceptible populations 7. To overcome this, researchers have often resorted to using a combination of antibiotics 8-10. Antibiotic resistance on a single species level can be either intrinsic or acquired. Intrinsic resistance is a characteristic that is coded for in the genome of the strain, independent of antibiotic pressure and not a consequence of horizontal gene transfer. An example of intrinsic resistance includes the outer membrane in Gram-negative bacteria, which modulates the cell permeability and limits the entry of hydrophobic antibiotics 11. Lacking an outer membrane, Gram-positive strains are generally less protected and antibiotics may enter these bacteria more easily 12. The result is that Gram-negative bacteria are less susceptible to many antibiotics that are effective against Gram-positive pathogens. A case in point is the bisindole antibiotic violacein, which is produced by a number of bacterial species 13 , including strains of Chromobacterium 14,15 , Janthinobacterium 16 , Collimonas 17 and Duganella 18. As an antibiotic, violacein is primarily active against Gram-positive strains 15,18-2...