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Background Burkholderia pseudomallei, a Gram-negative bacterium that causes melioidosis, was reported to produce biofilm. As the disease causes high relapse rate when compared to other bacterial infections, it therefore might be due to the reactivation of the biofilm forming bacteria which also provided resistance to antimicrobial agents. However, the mechanism on how biofilm can provide tolerance to antimicrobials is still unclear.Methodology/Principal FindingsThe change in resistance of B. pseudomallei to doxycycline, ceftazidime, imipenem, and trimethoprim/sulfamethoxazole during biofilm formation were measured as minimum biofilm elimination concentration (MBEC) in 50 soil and clinical isolates and also in capsule, flagellin, LPS and biofilm mutants. Almost all planktonic isolates were susceptible to all agents studied. In contrast, when they were grown in the condition that induced biofilm formation, they were markedly resistant to all antimicrobial agents even though the amount of biofilm production was not the same. The capsule and O-side chains of LPS mutants had no effect on biofilm formation whereas the flagellin-defective mutant markedly reduced in biofilm production. No alteration of LPS profiles was observed when susceptible form was changed to resistance. The higher amount of N-acyl homoserine lactones (AHLs) was detected in the high biofilm-producing isolates. Interestingly, the biofilm mutant which produced a very low amount of biofilm and was sensitive to antimicrobial agents significantly resisted those agents when grown in biofilm inducing condition.Conclusions/SignificanceThe possible drug resistance mechanism of biofilm mutants and other isolates is not by having biofilm but rather from some factors that up-regulated when biofilm formation genes were stimulated. The understanding of genes related to this situation may lead us to prevent B. pseudomallei biofilms leading to the relapse of melioidosis.

Transmission electron microscopy (TEM), scanning electron microscopy (SEM) and energy-dispersive analysis of X-ray (EDAX) have been used to examine chlorhexidine diacetate (CHA)-sensitive and -resistant isolates of Pseudomonas stutzeri and to determine the effects of CHA on the cells. Significant differences were observed in the structure, size and elemental composition of CHA-sensitive and -resistant cells. Treatment with CHA produced considerably greater changes in CHA-sensitive cells, with widespread peeling of the outer membrane, a substantial loss of cytoplasmic electron-dense material and extensive lysis. Cells from the resistant isolates showed no blebbing of the outer membrane and no structural damage. X-ray mapping confirmed the difference in CHA uptake between CHA-sensitive and CHA-resistant cells. It is proposed that changes in the outer membrane form a major mechanism of resistance to CHA in P. stutzeri.

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