Smith Kenneth P, Kirby James E
Department of Pathology, Beth Israel Deaconess Medical Center, Boston, Massachusetts, USA.
Department of Pathology, Beth Israel Deaconess Medical Center, Boston, Massachusetts, USA
J Clin Microbiol. 2016 Sep;54(9):2288-93. doi: 10.1128/JCM.00932-16. Epub 2016 Jun 22.
With rapid emergence of multidrug-resistant bacteria, there is often a need to perform susceptibility testing for less commonly used or newer antimicrobial agents. Such testing can often be performed only by using labor-intensive, manual dilution methods and lies outside the capacity of most clinical labs, necessitating reference laboratory testing and thereby delaying the availability of susceptibility data. To address the compelling clinical need for microbiology laboratories to perform such testing in-house, we explored a novel, automated, at-will broth microdilution-based susceptibility testing platform. Specifically, we used the modified inkjet printer technology in the HP D300 digital dispensing system to dispense, directly from stock solutions into a 384-well plate, the 2-fold serial dilution series required for broth microdilution testing. This technology was combined with automated absorbance readings and data analysis to determine MICs. Performance was verified by testing members of the Enterobacteriaceae for susceptibility to ampicillin, cefazolin, ciprofloxacin, colistin, gentamicin, meropenem, and tetracycline in comparison to the results obtained with a broth microdilution reference standard. In precision studies, essential and categorical agreement levels were 96.8% and 98.3%, respectively. Furthermore, significantly fewer D300-based measurements were outside ±1 dilution from the modal MIC, suggesting enhanced reproducibility. In accuracy studies performed using a panel of 80 curated clinical isolates, rates of essential and categorical agreement and very major, major, and minor errors were 94%, 96.6%, 0%, 0%, and 3.4%, respectively. Based on these promising initial results, it is anticipated that the D300-based methodology will enable hospital-based clinical microbiology laboratories to perform at-will broth microdilution testing of antimicrobials and to address a critical testing gap.
随着多重耐药菌的迅速出现，常常需要对较少使用的或新型抗菌药物进行药敏试验。此类试验通常只能通过劳动密集型的手工稀释方法进行，大多数临床实验室无法胜任，因此需要送交参考实验室检测，从而延迟了药敏数据的获取。为满足微生物实验室在内部开展此类检测的迫切临床需求，我们探索了一种基于肉汤微量稀释法的新型自动化、按需药敏检测平台。具体而言，我们在惠普D300数字分配系统中使用改良的喷墨打印机技术，将肉汤微量稀释试验所需的2倍系列稀释液直接从储备液分配到384孔板中。该技术与自动吸光度读数和数据分析相结合以确定最低抑菌浓度（MIC）。通过检测肠杆菌科细菌对氨苄西林、头孢唑林、环丙沙星、黏菌素、庆大霉素、美罗培南和四环素的敏感性，并与肉汤微量稀释参考标准获得的结果进行比较，验证了该平台的性能。在精密度研究中，基本一致率和绝对一致率分别为96.8%和98.3%。此外，基于D300的测量结果明显更少地超出模式MIC的±1个稀释度，表明重现性增强。在使用一组80株精选临床分离株进行的准确性研究中，基本一致率、绝对一致率以及极重大、重大和微小错误率分别为94%、96.6%、0%、0%和3.4%。基于这些令人鼓舞的初步结果，预计基于D300的方法将使医院临床微生物实验室能够开展抗菌药物的按需肉汤微量稀释试验，并填补关键的检测空白。
