Phenotypic characterization of multidrug-resistant Pseudomonas aeruginosa strains isolated from pediatric patients associated to biofilm formation.

BACKGROUND: Pseudomonas aeruginosa is an opportunistic pathogen that has acquired several mechanisms of resistance to multiple groups of antibiotic agents and has been widely employed as a model organism for the study of biofilm formation. Many P. aeruginosa structures embedded in the extracellular matrix, such as exopolysaccharides (EPS), flagella, and type-IV pili (T4P), have been associated with biofilm formation. In this study, we assess biofilm formation by crystal violet quantification in clinical strains of multidrug-resistant (MDR) P. aeruginosa isolated from the Hospital Infantil de México Federico Gómez (HIMFG) associated to total and reducing EPS production (quantification by the anthrone and DNS method, respectively), twitching motility activity by T4P, and flagellar-mediated motility.

RESULTS: The determination of Minimum Inhibitory Concentration (MIC) showed that >50% of P. aeruginosa strains were resistant to 12 different antibiotics (TIC, CAZ, CTX, CRO, FEP, AZT, GM, CIP, LEV, PZT, IMP, and MEM). Total and reducing EPS analysis of the 58 biofilm-forming MDR P. aeruginosa strains showed heterogeneous values ranging from OD600 9.06 to 212.33, displaying a linear correlation with the production of total EPS (59.66μg/ml to 6000.33μg/ml; R(2)=0.89), and a higher correlation with reducing EPS (88.33μg/ml to 1100.66μg/ml; R(2)=0.96). T4P twitching motility showed a moderated linear correlation (2.00mm to 28.33mm; R(2)=0.74). Even though it has been demonstrated that flagella contribute to the initial stages of biofilm formation, crystal violet analysis showed a moderate correlation (R(2)=0.49) with flagellar-mediated motility in MDR P. aeruginosa under the tested conditions. In addition, PFGE profiles revealed two subgroups generating profiles group A, consisting of 89.63% (52/58) of the strains, and group B, consisting of 13.09% (6/58) of the strains.

CONCLUSIONS: Phenotypic analysis showed a correlation among the biofilms developed in the MDR P. aeruginosa strains with EPS (total and reducing) production, T4P-activity by twitching motility and flagellar-mediated motility.