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首页> 外文期刊>Journal of Clinical Microbiology >Direct Identification of Bacteria from Positive Blood Cultures by Amplification and Sequencing of the 16S rRNA Gene: Evaluation of BACTEC 9240 Instrument True- Positive and False-Positive Results
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Direct Identification of Bacteria from Positive Blood Cultures by Amplification and Sequencing of the 16S rRNA Gene: Evaluation of BACTEC 9240 Instrument True- Positive and False-Positive Results

机译：通过扩增和16S rRNA基因测序直接从阳性血液培养物中鉴定细菌：BACTEC 9240仪器真假阳性结果的评估

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In a previous study which evaluated the BACTEC 9240 automated blood culture system (Becton Dickinson Diagnostic Instrument Systems, Sparks, Md.), we noted a 1.3% “instrument false-positive” rate. That is, the BACTEC system signaled that a bottle (BACTEC Plus Aerobic/F bottle or BACTEC Anaerobic Lytic/10 bottle) culture was positive but a Gram stain was negative and there was no growth of bacteria or yeasts on subculture to chocolate agar. Furthermore, from the same sample of blood, cultures for fungi using the Isolator blood culture system (Wampole Laboratories, Cranbury, N.J.) were negative for growth. For the present study, we evaluated 76 instrument false-positive samples for the presence of 16S ribosomal DNA using the MicroSeq 500 kit (PE Biosystems, Foster City, Calif.). These samples also were negative for fungi by the Isolator method. This kit has a PCR module and sequencing module for the amplification and sequencing of the 16S RNA gene and provides a database for sequence alignment and identification of bacteria. To optimize the assay, we evaluated the effect of adding 0.5% bovine serum albumin to the sample from blood culture bottles and found that it decreased the effects of inhibitors on the PCR. Two control groups of blood culture specimens were also evaluated. One group (n = 45) were “instrument true positives”; the instrument signaled positive, and subsequent Gram stains were positive and subcultures on chocolate agar grew bacteria. The other group (n = 20) were “instrument true negatives”; the instrument signaled negative, the Gram stain was negative, and subcultures on chocolate agar and from the Isolator tube on fungal media showed no growth. None of the 76 instrument false-positive samples had evidence for 16S rRNA gene sequences. All of the instrument true-positive samples and all of the instrument true-negative specimens were positive and negative, respectively, using the MicroSeq 500 kit. Total peripheral white blood cell counts were statistically significantly higher for patients who had instrument false-positive results than for patients who had instrument true-positive or true-negative results (P = 0.001). We conclude that instrument false positives signaled by the BACTEC 9240 system are not due to bacteria in the blood culture samples.
机译：在先前的评估BACTEC 9240自动血液培养系统（马里兰州Sparks的Becton Dickinson诊断仪器系统）的研究中，我们注意到1.3％的“仪器假阳性”率。也就是说，BACTEC系统发出信号，表明一个瓶（BACTEC Plus有氧/ F瓶或BACTEC厌氧Lytic / 10瓶）培养物为阳性，但革兰氏染色为阴性，继代培养到巧克力琼脂上没有细菌或酵母的生长。此外，从相同的血液样本中，使用Isolator血液培养系统（Wampole Laboratories，Cranbury，N.J。）的真菌培养物生长阴性。在本研究中，我们使用MicroSeq 500试剂盒（PE Biosystems，Foster City，CA）评估了76种仪器假阳性样品中16S核糖体DNA的存在。这些样品也通过隔离器方法阴性。该试剂盒具有用于扩增和测序16S RNA基因的PCR模块和测序模块，并提供了用于序列比对和细菌鉴定的数据库。为了优化测定，我们评估了向血液培养瓶中的样品中添加0.5％牛血清白蛋白的效果，并发现其降低了抑制剂对PCR的影响。还评估了两个对照组的血液培养标本。一组（ n = 45）为“仪器真实阳性”;仪器发出阳性信号，随后的革兰氏染色阳性，并且在巧克力琼脂上继代培养细菌。另一组（ n = 20）是“仪器真阴性”;仪器指示为阴性，革兰氏染色为阴性，在巧克力琼脂上以及在真菌培养基上的隔离管中的亚培养物均未见生长。 76个仪器假阳性样品中没有一个具有16S rRNA基因序列的证据。使用MicroSeq 500试剂盒，所有仪器真阳性样品和所有仪器真阴性样品分别为阳性和阴性。具有仪器假阳性结果的患者的外周血白细胞总数明显高于具有仪器真阳性或真阴性结果的患者（ P = 0.001）。我们得出的结论是，BACTEC 9240系统发出的仪器误报并不是由于血液培养样本中的细菌引起的。

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