DNase I and chitosan enhance efficacy of ceftazidime to eradicate Burkholderia pseudomallei biofilm cells.

Biofilm-associated Burkholderia pseudomallei infection contributes to antibiotic resistance and relapse of melioidosis. Burkholderia pseudomallei biofilm matrix contains extracellular DNA (eDNA) that is crucial for biofilm establishment. However, the contribution of eDNA to antibiotic resistance by B. pseudomallei remains unclear. In this study, we first demonstrated in vitro that DNase I with the administration of ceftazidime (CAZ) at 24 h considerably inhibited the 2-day biofilm formation and reduced the number of viable biofilm cells of clinical B. pseudomallei isolates compared to biofilm treated with CAZ alone. A 3-4 log reduction in numbers of viable cells embedded in the 2-day biofilm was observed when CAZ was combined with DNase I. Confocal laser-scanning microscope visualization emphasized the competence of DNase I followed by CAZ supplementation to significantly limit B. pseudomallei biofilm development and to eradicate viable embedded B. pseudomallei biofilm cells. Furthermore, DNase I supplemented with chitosan (CS) linked with CAZ (CS/CAZ) significantly eradicated shedding planktonic and biofilm cells. These findings indicated that DNase I effectively degraded eDNA leading to biofilm inhibition and dispersion, subsequently allowing CAZ and CS/CAZ to eradicate both shedding planktonic and embedded biofilm cells. These findings provide efficient strategies to interrupt biofilm formation and improve antibiotic susceptibility of biofilm-associated infections.

Chitosan biological molecule improves bactericidal competence of ceftazidime against Burkholderia pseudomallei biofilms.

Biofilm-associated Burkholderia pseudomallei infections (melioidosis) are problematic because of reduced sensitivity to antibiotics and high frequency of relapse. Biofilm dispersal agents are essential to liberate the biofilm-encased cells, which then become planktonic and are more susceptible to antibiotics. This study aimed to evaluate the ability of deacetylated chitosan (dCS), an antimicrobial and antibiofilm biological macromolecule, to disrupt established biofilms, thus enabling ceftazidime (CAZ) to kill biofilm-embedded B. pseudomallei. We combined dCS with CAZ using a mechanical stirring method to generate dCS/CAZ. In combination, 1.25-2.5 mg ml-1 dCS/1-2 µg ml-1 CAZ acted synergistically to kill cells more effectively than did either dCS or CAZ alone. Notably, a combination of 5-10 mg ml-1 dCS with 256-512 µg ml-1 CAZ, prepared either by mechanical stirring (dCS/CAZ) or mixing (dCS + CAZ), drastically improved bactericidal activities against biofilm cells leading to a 3-6 log CFU reduction. Confocal laser-scanning microscope (CLSM) images revealed that 10 mg ml-1 dCS/512 µg ml-1 CAZ is by far the best formulation to diminish B. pseudomallei biofilm biomass and produces the lowest live/dead cell ratios of B. pseudomallei in biofilm matrix. Collectively, these findings emphasize the potential of novel therapeutic antibacterial and antibiofilm agents to fight against antibiotic-tolerant B. pseudomallei biofilm-associated infections.