Calderwood S B, Baker M A, Carroll P A, Michel J L, Arbeit R D, Ausubel F M
Infectious Disease Unit, Massachusetts General Hospital, Boston 02114, USA.
J Clin Microbiol. 1996 Nov;34(11):2860-5. doi: 10.1128/jcm.34.11.2860-2865.1996.
We examined the utility of a PCR-based method termed cleaved amplified polymorphic sequences (CAPS) to type 35 well-characterized isolates of Staphylococcus epidermidis. The results were compared with detailed epidemiologic information and typing obtained by using pulsed-field gel electrophoresis (PFGE). To identify CAPS markers for this study, eight pairs of oligonucleotide primers corresponding to five previously sequenced S. epidermidis genes were synthesized and then used to amplify DNA sequences from the S. epidermidis strains by using PCR. Amplified products were reproducibly obtained for seven of eight primer pairs from chromosomal DNA of 33 of the 35 isolates. Seven restriction site polymorphisms were found in five of the amplified products when they were subjected to digestion with a panel of restriction endonucleases. Each fragment-enzyme combination that was polymorphic demonstrated only two alleles in the 33 S. epidermidis isolates analyzed, corresponding to the presence or absence of a single restriction site. Overall, five distinct combinations of alleles were detected and were designated CAPS types A through E. There was a close correlation between the CAPS grouping, the epidemiologic information for the strains, and grouping by PFGE following SmaI digestion of chromosomal DNA. Although PFGE analysis was more discriminatory than typing based on the limited number of CAPS markers used in this study (isolates from the same CAPS group were sometimes distributed into more than one PFGE group), no isolates from the same PFGE group were found in more than one CAPS group. The CAPS procedure was highly reproducible, in contrast to published experience with arbitrarily primed PCR. These preliminary data suggest that CAPS represents a PCR-based technique for strain typing that is highly reproducible, rapid, utilizes widely available technologies, and provides results that are relatively easy to interpret and express.
我们检测了一种基于聚合酶链反应(PCR)的方法——酶切扩增多态性序列(CAPS),用于对35株特征明确的表皮葡萄球菌进行分型。将结果与详细的流行病学信息以及使用脉冲场凝胶电泳(PFGE)获得的分型结果进行比较。为了确定本研究的CAPS标记,合成了对应于五个先前已测序的表皮葡萄球菌基因的八对寡核苷酸引物,然后通过PCR用于扩增表皮葡萄球菌菌株的DNA序列。从35株分离株中的33株的染色体DNA中,八对引物中的七对可重复获得扩增产物。当用一组限制性内切酶对五个扩增产物进行酶切时,发现了七个限制性位点多态性。在分析的33株表皮葡萄球菌分离株中,每个具有多态性的片段 - 酶组合仅显示两个等位基因,对应于单个限制性位点的存在或不存在。总体而言,检测到五个不同的等位基因组合,并命名为CAPS A型至E型。CAPS分组、菌株的流行病学信息以及染色体DNA经SmaI酶切后的PFGE分组之间存在密切相关性。虽然PFGE分析比基于本研究中使用的有限数量的CAPS标记的分型更具鉴别力(来自同一CAPS组的分离株有时分布在多个PFGE组中),但未发现来自同一PFGE组的分离株存在于多个CAPS组中。与已发表的任意引物PCR经验相反,CAPS程序具有高度可重复性。这些初步数据表明,CAPS代表了一种基于PCR的菌株分型技术,该技术具有高度可重复性、快速、利用广泛可用的技术,并且提供相对易于解释和表达的结果。