Laboratorio de Investigación en Bacteriología Intestinal, Unidad de Hemato-Onocología e Investigación. Hospital Infantil de México Federico Gómez, Dr. Márquez 162, Col. Doctores, Delegación Cuauhtémoc, México, D.F. 06720, Mexico.
Laboratorio de Infectología, Departamento de Infectología. Hospital Infantil de México Federico Gómez, Dr. Márquez 162, Col. Doctores, Delegación Cuauhtémoc, México, D.F. 06720, Mexico.
Microbiol Res. 2015 Mar;172:68-78. doi: 10.1016/j.micres.2014.11.005. Epub 2014 Dec 5.
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.
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.
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.
铜绿假单胞菌是一种机会性病原体，已经获得了多种机制来抵抗多组抗生素，并被广泛用作生物膜形成研究的模式生物。许多嵌入细胞外基质的铜绿假单胞菌结构，如胞外多糖（EPS）、鞭毛和 IV 型菌毛（T4P），与生物膜形成有关。在这项研究中，我们通过结晶紫定量评估了从墨西哥 Federico Gómez 儿童医院（HIMFG）分离的多药耐药（MDR）铜绿假单胞菌临床株的生物膜形成，这些临床株与总 EPS 和还原 EPS 的产生（分别通过蒽酮和 DNS 法定量）、T4P 介导的翻滚运动活性以及鞭毛介导的运动有关。
最低抑菌浓度（MIC）的测定表明，超过 50%的铜绿假单胞菌菌株对 12 种不同的抗生素（TIC、CAZ、CTX、CRO、FEP、AZT、GM、CIP、LEV、PZT、IMP 和 MEM）有耐药性。对 58 株生物膜形成的 MDR 铜绿假单胞菌的总 EPS 和还原 EPS 分析显示，值从 OD600 9.06 到 212.33 不等，与总 EPS 的产生呈线性相关（59.66μg/ml 至 6000.33μg/ml；R(2)=0.89），与还原 EPS 的相关性更高（88.33μg/ml 至 1100.66μg/ml；R(2)=0.96）。T4P 翻滚运动显示出适度的线性相关性（2.00mm 至 28.33mm；R(2)=0.74）。尽管已经证明鞭毛有助于生物膜形成的初始阶段，但在测试条件下，结晶紫分析显示与 MDR 铜绿假单胞菌的鞭毛介导运动呈中度相关（R(2)=0.49）。此外，PFGE 图谱显示了两个亚群，产生图谱 A，由 89.63%（52/58）的菌株组成，图谱 B，由 13.09%（6/58）的菌株组成。
表型分析显示，MDR 铜绿假单胞菌菌株的生物膜形成与 EPS（总 EPS 和还原 EPS）产生、T4P 活性（翻滚运动）和鞭毛介导的运动之间存在相关性。
