Snelling A M, Gerner-Smidt P, Hawkey P M, Heritage J, Parnell P, Porter C, Bodenham A R, Inglis T
Department of Microbiology, University of Leeds, United Kingdom.
J Clin Microbiol. 1996 May;34(5):1193-202. doi: 10.1128/jcm.34.5.1193-1202.1996.
Acinetobacter spp. are being reported with increasing frequency as causes of nosocomial infection. In order to identify reservoirs of infection as quickly as possible, a rapid typing method that can differentiate epidemic strains from environmental and nonepidemic strains is needed. In 1993, a cluster of Acinetobacter baumannii isolates from five patients in the adult intensive therapy unit of our tertiary-care teaching hospital led us to develop and optimize a rapid repetitive extragenic palindromic sequence-based PCR (REP-PCR) typing protocol for members of the Acinetobacter calcoaceticus-A. baumannii complex that uses boiled colonies and consensus primers aimed at repetitive extragenic palindromic sequences. Four of the five patient isolates gave the same REP-PCR typing pattern as isolates of A. baumannii obtained from the temperature probe of a Bennett humidifier; the fifth isolate had a unique profile. Disinfection of the probe with 70% ethanol, as recommended by the manufacturer, proved ineffective, as A. baumannii with the same REP-PCR pattern was isolated from it 10 days after cleaning, necessitating a change in our decontamination procedure. Results obtained with REP-PCR were subsequently confirmed by ribotyping. To evaluate the discriminatory power (D) of REP-PCR for typing members of the A. calcoaceticus-A. baumannii complex, compared with that of ribotyping, we have applied both methods to a collection of 85 strains that included representatives of six DNA groups within the complex. Ribotyping using EcoRI digests yielded 53 patterns (D = 0.98), whereas 68 different REP-PCR patterns were observed (D = 0.99). By computer-assisted analysis of gel images, 74 patterns were observed with REP-PCR (D = 1.0). Overall, REP-PCR typing proved to be slightly more discriminatory than ribotyping. Our results indicate that REP-PCR typing used boiled colonies is a simple, rapid, and effective means of typing members of the A. calcoaceticus-A. baumannii complex.
不动杆菌属作为医院感染的病因，其报告频率日益增加。为了尽快确定感染源，需要一种能够区分流行菌株与环境菌株和非流行菌株的快速分型方法。1993年，我们三级护理教学医院成人重症监护病房的5名患者中分离出的一群鲍曼不动杆菌，促使我们开发并优化了一种基于快速重复基因外回文序列的PCR（REP-PCR）分型方案，用于醋酸钙不动杆菌-鲍曼不动杆菌复合体的成员，该方案使用煮沸的菌落和针对重复基因外回文序列的通用引物。5名患者分离株中的4株与从贝内特加湿器温度探头获得的鲍曼不动杆菌分离株具有相同的REP-PCR分型模式；第5株分离株具有独特的图谱。按照制造商的建议，用70%乙醇对探头进行消毒，结果证明无效，因为在清洁10天后，仍从探头中分离出具有相同REP-PCR模式的鲍曼不动杆菌，这就需要改变我们的去污程序。REP-PCR的结果随后通过核糖体分型得到证实。为了评估REP-PCR对醋酸钙不动杆菌-鲍曼不动杆菌复合体成员分型的鉴别力（D），并与核糖体分型进行比较，我们将这两种方法应用于一组85株菌株，其中包括该复合体内6个DNA组的代表菌株。使用EcoRI酶切的核糖体分型产生了53种图谱（D = 0.98），而观察到68种不同的REP-PCR图谱（D = 0.99）。通过对凝胶图像的计算机辅助分析，REP-PCR观察到74种图谱（D = 1.0）。总体而言，REP-PCR分型的鉴别力略高于核糖体分型。我们的结果表明，使用煮沸菌落的REP-PCR分型是醋酸钙不动杆菌-鲍曼不动杆菌复合体成员分型的一种简单、快速且有效的方法。
