红霉素耐药决定因素的微阵列分析。

Microarray analysis of erythromycin resistance determinants.

作者信息

Volokhov D, Chizhikov V, Chumakov K, Rasooly A

机构信息

FDA Center for Food Safety and Applied Nutrition, College Park, MD 20740-3835, USA.

出版信息

J Appl Microbiol. 2003;95(4):787-98. doi: 10.1046/j.1365-2672.2003.02046.x.

DOI:10.1046/j.1365-2672.2003.02046.x

PMID:12969293

Abstract

AIMS

To develop a DNA microarray for analysis of genes encoding resistance determinants to erythromycin and the related macrolide, lincosamide and streptogramin B (MLS) compounds.

METHODS AND RESULTS

We developed an oligonucleotide microarray containing seven oligonucleotide probes (oligoprobes) for each of the six genes (ermA, ermB, ermC, ereA, ereB and msrA/B) that account for more than 98% of MLS resistance in Staphylococcus aureus clinical isolates. The microarray was used to test reference and clinical S. aureus and Streptococcus pyrogenes strains. Target genes from clinical strains were amplified and fluorescently labelled using multiplex PCR target amplification. The microarray assay correctly identified the MLS resistance genes in the reference strains and clinical isolates of S. aureus, and the results were confirmed by direct DNA sequence analysis. Of 18 S. aureus clinical strains tested, 11 isolates carry MLS determinants. One gene (ermC) was found in all 11 clinical isolates tested, and two others, ermA and msrA/B, were found in five or more isolates. Indeed, eight (72%) of 11 clinical isolate strains contained two or three MLS resistance genes, in one of the three combinations (ermA with ermC, ermC with msrA/B, ermA with ermC and msrA/B).

CONCLUSIONS

Oligonucleotide microarray can detect and identify the six MLS resistance determinants analysed in this study.

SIGNIFICANCE AND IMPACT OF THE STUDY

Our results suggest that microarray-based detection of microbial antibiotic resistance genes might be a useful tool for identifying antibiotic resistance determinants in a wide range of bacterial strains, given the high homology among microbial MLS resistance genes.

摘要

目的

开发一种DNA微阵列，用于分析编码对红霉素及相关大环内酯类、林可酰胺类和链阳菌素B（MLS）化合物耐药决定簇的基因。

方法与结果

我们开发了一种寡核苷酸微阵列，针对在金黄色葡萄球菌临床分离株中占MLS耐药性98%以上的六个基因（ermA、ermB、ermC、ereA、ereB和msrA/B），每个基因包含七个寡核苷酸探针（寡探针）。该微阵列用于检测参考菌株和临床金黄色葡萄球菌及化脓性链球菌菌株。使用多重PCR靶标扩增对临床菌株的靶基因进行扩增和荧光标记。微阵列分析正确鉴定了金黄色葡萄球菌参考菌株和临床分离株中的MLS耐药基因，并通过直接DNA序列分析证实了结果。在测试的18株金黄色葡萄球菌临床菌株中，11株分离株携带MLS决定簇。在所测试的所有11株临床分离株中均发现一个基因（ermC），另外两个基因ermA和msrA/B在五株或更多分离株中被发现。实际上，11株临床分离株中有八株（72%）包含两到三个MLS耐药基因，呈三种组合之一（ermA与ermC、ermC与msrA/B、ermA与ermC和msrA/B）。