van Belkum A, Kluytmans J, van Leeuwen W, Bax R, Quint W, Peters E, Fluit A, Vandenbroucke-Grauls C, van den Brule A, Koeleman H
Department of Bacteriology, University Hospital Dijkzigt, Rotterdam, The Netherlands.
J Clin Microbiol. 1995 Jun;33(6):1537-47. doi: 10.1128/jcm.33.6.1537-1547.1995.
Fifty-nine isolates of Staphylococcus aureus and a single strain of Staphylococcus intermedius were typed by arbitrarily primed PCR (AP-PCR). To study reproducibility and discriminatory abilities, AP-PCR was carried out in seven laboratories with a standardized amplification protocol, template DNA isolated in a single institution, and a common set of three primers with different resolving powers. The 60 strains could be divided into 16 to 30 different genetic types, depending on the laboratory. This difference in resolution was due to differences in technical procedures (as shown by the deliberate introduction of experimental variables) and/or the interpretation of the DNA fingerprints. However, this did not hamper the epidemiologically correct clustering of related strains. The average number of different genotypes identified exceeded those of the more traditional typing strategies (F. C. Tenover, R. Arbeit, G. Archer, J. Biddle, S. Byrne, R. Goering, G. Hancock, G. A. Hebert, B. Hill, R. Hollis, W. R. Jarvis, B. Kreiswirth, W. Eisner, J. Maslow, L. K. McDougal, J. M. Miller, M. Mulligan, and M. A. Pfaller, J. Clin. Microbiol. 32:407-415, 1994). Comparison of AP-PCR with pulsed-field gel electrophoresis (PFGE) indicated the existence of strains with constant PFGE types but variable AP-PCR types. The reverse (constant AP-PCR and variable PFGE patterns) was also observed. This indicates additional resolution for combined analyses. It is concluded that AP-PCR is well suited for genetic analysis and monitoring of nosocomial spreading of staphylococci. The interlaboratory reproducibility of DNA-banding patterns and the intralaboratory standardization need improvement.
采用随机引物PCR(AP-PCR)技术对59株金黄色葡萄球菌和1株中间葡萄球菌进行分型。为研究其重复性和鉴别能力,在7个实验室按照标准化扩增方案进行AP-PCR实验,模板DNA来自同一机构,使用一组具有不同分辨能力的通用引物。根据实验室的不同,这60株菌株可分为16至30种不同的基因类型。这种分辨率的差异是由于技术操作的不同(如故意引入实验变量所示)和/或对DNA指纹图谱的解读不同。然而,这并不妨碍相关菌株在流行病学上的正确聚类。所鉴定的不同基因型的平均数量超过了更传统的分型策略(F.C. Tenover、R. Arbeit、G. Archer、J. Biddle、S. Byrne、R. Goering、G. Hancock、G. A. Hebert、B. Hill、R. Hollis、W. R. Jarvis、B. Kreiswirth、W. Eisner、J. Maslow、L. K. McDougal、J. M. Miller、M. Mulligan和M. A. Pfaller,《临床微生物学杂志》32:407 - 415,1994年)。AP-PCR与脉冲场凝胶电泳(PFGE)的比较表明,存在PFGE类型恒定但AP-PCR类型可变的菌株。也观察到了相反的情况(AP-PCR类型恒定而PFGE模式可变)。这表明联合分析具有额外的分辨率。得出的结论是,AP-PCR非常适合用于葡萄球菌医院内传播的基因分析和监测。DNA条带模式的实验室间重复性和实验室内标准化需要改进。