A novel species-specific PCR assay for identifying Lactobacillus fermentum
A novel species-specific PCR assay for identifying Lactobacillus fermentum
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DOI：
10.1099/jmm.0.45770-0
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发表时间：
2005-03-01
影响因子：
3
通讯作者：
Macpherson, L
中科院分区：
文献类型：
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作者：
Dickson, EM;Riggio, MP;Macpherson, L
发酵乳杆菌是一种革兰氏阳性细菌，与活动性龋齿病变有关。传统上鉴定乳酸菌种类的方法是基于培养方法加上生化试验，这通常是不可靠的。本研究的目的是建立一种直接检测口腔临床样品中发酵乳杆菌的物种特异性PCR方法。通过对细菌16S rRNA基因进行比对，并在其3'端选择L. fermentum特异性序列，确定了L. fermentum特异性PCR引物。扩增产物长度为337 bp，表明发酵乳杆菌DNA PCR阳性。结果表明，引物对发酵乳杆菌DNA具有特异性，因为当使用其他多种口腔细菌（包括密切相关的乳酸菌）的基因组DNA作为测试物种时，没有获得PCR产物。PCR检测随后被用于鉴定龈上菌斑样本和急性牙槽脓肿患者吸出的脓液中的发酵乳杆菌DNA。70个（5.7%）龈上菌斑样本中有4个检测到发酵乳杆菌DNA阳性，而19个脓液样本中没有一个检测到发酵乳杆菌DNA阳性。在临床标本中，该PCR检测提供了一种比传统培养方法更快速、特异和敏感的方法来鉴定发酵乳杆菌。
Lactobacillus fermentum is a Gram-positive bacterium that is associated with active caries lesions. Methods for identifying Lactobacillus species traditionally have been based upon culture methods coupled with biochemical tests, which are generally unreliable. The aim of this study was to develop a species-specific PCR assay for the direct detection of L. fermentum in oral clinical samples. PCR primers specific for L. fermentum were identified by alignment of bacterial 16S rRNA genes and selection of sequences specific for L. fermentum at their 3' ends. PCR positivity for L. fermentum DNA was indicated by amplification of a 337 bp product. The primers were shown to be specific for L. fermentum DNA, since no PCR product was obtained when genomic DNA from a wide range of other oral bacteria, including closely related Lactobacillus species, were used as test species. The PCR assay was then used in an attempt to identify L. fermentum DNA in supragingival plaque samples and in pus aspirates from subjects with acute dento-alveolar abscesses. Four out of 70 (5(.)7%) supragingival plaque samples analysed were positive for the presence of L. fermentum, DNA while none of the 19 pus samples analysed was positive for L. fermentum DNA. This PCR assay provides a more rapid, specific and sensitive alternative to conventional culture methods for the identification of L. fermentum, in clinical specimens.