Maciá Maria D, Borrell Nuria, Pérez José L, Oliver Antonio
Servicio de Microbiología, Hospital Son Dureta, C. Andrea Doria No. 55, 07014 Palma de Mallorca, Spain.
Antimicrob Agents Chemother. 2004 Jul;48(7):2665-72. doi: 10.1128/AAC.48.7.2665-2672.2004.
Resistance development in Pseudomonas aeruginosa from chronically colonized cystic fibrosis (CF) patients has been linked to the presence of a high proportion of mismatch repair-deficient hypermutable strains. The detection of hypermutable strains by microbiology laboratories may be useful for establishing adequate antimicrobial therapies. In this work, we find that the Etest and disk diffusion can be used as simple methods for the detection and susceptibility testing of hypermutable P. aeruginosa isolates. Strain PAO1 and its hypermutable derivative strain PAODeltamutS were used to standardize the procedure, which was tested with 35 P. aeruginosa isolates from 21 CF patients. Mutation frequencies were estimated by standard methods, and 29% of the isolates were found to be hypermutable. MICs and inhibition zone diameters were determined for ceftazidime, imipenem, meropenem, ciprofloxacin, and tobramycin by using Etest strips and conventional disks, respectively. The presence (or absence) of resistant mutant subpopulations, as well as their relative numbers and the highest MICs for them (or smallest inhibition zone diameters), was recorded. The presence of resistant mutant subpopulations within the inhibition zones of three or more antibiotics clearly identified the strains as hypermutable (they were present in 10 of 10 hypermutable strains and 0 of 25 nonhypermutable strains) with both methods. Additionally, these methods allowed us to differentiate between dual effects of hypermutation in antibiotic resistance, namely, that (i) hypermutable isolates were substantially more resistant than nonhypermutable isolates and that (ii) the resistance of hypermutable isolates was dramatically increased by the presence of resistant mutant subpopulations. This differentiation may be relevant for the design of adequate treatments, since the second effect, in contrast to the first, may be overcome by antibiotic combinations.
来自长期定植的囊性纤维化（CF）患者的铜绿假单胞菌耐药性的产生与高比例错配修复缺陷的超突变菌株的存在有关。微生物实验室检测超突变菌株可能有助于确定适当的抗菌治疗方法。在这项研究中，我们发现Etest和纸片扩散法可作为检测超突变铜绿假单胞菌分离株及其药敏试验的简单方法。使用菌株PAO1及其超突变衍生菌株PAODeltamutS对该方法进行标准化，并对来自21例CF患者的35株铜绿假单胞菌分离株进行了测试。通过标准方法估计突变频率，发现29%的分离株为超突变株。分别使用Etest试纸条和传统纸片测定了头孢他啶、亚胺培南、美罗培南、环丙沙星和妥布霉素的MIC和抑菌圈直径。记录耐药突变亚群的存在（或不存在）、其相对数量以及它们的最高MIC（或最小抑菌圈直径）。两种方法均显示，在三种或更多抗生素的抑菌圈内存在耐药突变亚群可明确鉴定这些菌株为超突变株（10株超突变株中有10株存在，25株非超突变株中无）。此外，这些方法使我们能够区分超突变在抗生素耐药性中的双重作用，即：（i）超突变分离株比非超突变分离株耐药性更强；（ii）耐药突变亚群的存在使超突变分离株的耐药性显著增加。这种区分可能与设计适当的治疗方法相关，因为与第一种作用相反，第二种作用可能可通过联合使用抗生素来克服。
