Development of a multiplex-PCR for direct detection of the genes for enterotoxin B and C, and toxic shock syndrome toxin-1 in Staphylococcus aureus isolates.
Development of a multiplex-PCR for direct detection of the genes for enterotoxin B and C, and toxic shock syndrome toxin-1 in Staphylococcus aureus isolates.
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开发多重 PCR 直接检测金黄色葡萄球菌分离株中肠毒素 B 和 C 以及中毒性休克综合征毒素 1 的基因。
DOI：
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发表时间：
1998
影响因子：
3
通讯作者：
K. Köhrer
中科院分区：
文献类型：
--
作者：
F. Schmitz;M. Steiert;B. Hofmann;J. Verhoef;U. Hadding;H. Heinz;K. Köhrer
除了免疫扩散、酶联免疫吸附试验或凝集等检测金黄色葡萄球菌毒素产生的传统方法外，像聚合酶链式反应这样的扩增技术也可以非常敏感和特异地识别与肠毒素B和C以及TSST-1产生有关的基因。这些毒素可能是中毒性休克综合征(TSS)的原因之一。因此，需要一种简单、快速的检测系统来确定金黄色葡萄球菌分离株的毒素产生模式，以便可以更快、更容易地识别出疑似产生毒素的菌株。在本研究中，使用了一种新的多重-PCR方法，该方法允许在琼脂平板上生长的单个细菌菌落直接用于PCR检测，而不需要预先准备。该程序在一次测试中在4小时内产生关于SEB、SEC-1和TST基因存在的信息。为了分析该方法的敏感性和特异性，对100株耐甲氧西林金黄色葡萄球菌(MRSA)、50株凝固酶阴性葡萄球菌和50株其他真细菌进行了初步检测。将该扩增技术与一种常用的毒素检测方法--反向被动胶乳凝集法(RPLA)进行了比较。以Rpla检测结果为依据，SEB和TST的敏感度和特异度均为100%，而SEC-1的敏感度和特异度分别为100%和82%。利用SEC-1引物组，两个分离物被鉴定为携带相应的毒素基因，尽管RLA试验没有显示任何可检测到的毒素。多重-聚合酶链式反应快速产生有关葡萄球菌菌株产毒能力的可靠信息，并且可以很容易地整合到前面描述的多重程序中。后者使鉴定真细菌和葡萄球菌的特定聚合酶链式反应产物以及检测CoA和mecA基因成为可能。
As well as conventional methods such as immunodiffusion, ELISA, or agglutination for the detection of toxin production in Staphylococcus aureus, amplification techniques like PCR allow a very sensitive and specific identification of the genes responsible for enterotoxin B and C, and TSST-1 production. These toxins might be a cause of the toxic shock syndrome (TSS). For that reason an easy and quick test system for determining the toxin production pattern of S. aureus isolates is desirable so that strains suspected to be toxin producers may be identified much faster and easier. In the present investigation, a new multiplex-PCR method was used that allowed single bacterial colonies grown on agar plates to be used directly in the PCR assay without preceding preparation. This procedure generated information concerning the presence of seb, sec-1 and tst genes within 4 h in a single test. To analyse the sensitivity and the specificity of this procedure, 100 methicillin-resistant S. aureus (MRSA), 50 coagulase-negative staphylococci and 50 other eubacterial isolates were tested initially with sets of single primer pairs followed by a combined multiplex-PCR. Results of this amplification technique were compared to a conventional and widely used method for toxin detection, reversed passive latex agglutination (RPLA). With the RPLA assay results as the basis, sensitivity and specificity of the seb and tst primer sets were 100%, whereas sensitivity and specificity of the sec-1 primer set were 100% and 82%, respectively. With the sec-1 primer set, two isolates were identified as carrying the corresponding toxin gene although the RPLA test did not show any detectable toxin. The multiplex-PCR rapidly generated reliable information concerning the toxin-producing capacity of staphylococcal strains and could be easily integrated into a multiplex procedure described previously. The latter enabled the identification of specific PCR products for eubacteria and staphylococci as well as the detection of the coa and mecA genes.
中毒性休克综合征毒素1的核苷酸和部分氨基酸序列。
DOI：
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发表时间：
1986
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Blomster-Hautamaa,DA;Kreiswirth,BN;Kornblum,JS;Novick,RP;Schlievert,PM
通讯作者：
Schlievert,PM
大肠杆菌和金黄色葡萄球菌中葡萄球菌肠毒素 B 基因的分子克隆。
DOI：
10.1073/pnas.82.17.5850
发表时间：
1985
影响因子：
11.1
作者：
Ranelli,DM;Jones,CL;Johns,MB;Mussey,GJ;Khan,SA
通讯作者：
Khan,SA