Bennett Victoria M, McNevin Dennis, Roffey Paul, Gahan Michelle E
National Centre for Forensic Studies, Faculty of Education, Science, Technology and Mathematics, University of Canberra, Bruce, ACT, 2617, Australia.
Specialist Operations-Forensics, Australian Federal Police, PO Box 401, Canberra, ACT, 2601, Australia.
Forensic Sci Med Pathol. 2017 Mar;13(1):10-19. doi: 10.1007/s12024-016-9824-7. Epub 2016 Dec 26.
Yersinia pestis is a biological agent of high risk to national security due to its ability to be easily disseminated and transmitted among humans. If Y. pestis was to be utilized in a deliberate disease outbreak it would be essential to rapidly and accurately identify the agent. Current identification methods for Yersinia species are limited by their reliance on cultivation, the time taken to achieve results and/or the use of protocols that are not amenable for field use. Faster identification methods are urgently required. Microfluidic capillary electrophoresis was used to identify seven Yersinia species based on their protein profiles. Further objectives included determining if Yersinia species could be detected in mixtures of milk products and Escherichia coli, determining if Yersinia could be detected in a blinded identification and reproducibility across two platforms. Two characteristic protein bands were detected at 50 kilodaltons (kDa) and between 50 and 75 kDa for the Yersinia species. Individual Yersinia species could be differentiated from one another and distinguished from E. coli, Bacillus anthracis Sterne strain and Dipel (containing Bacillus thuringiensis). Due to the high protein content of milk products Yersinia could not be detected when mixed with these but was detected when mixed with E. coli. Species were correctly identified with 96% success in blinded procedures using 12 individuals. Whilst protein profile patterns were reproducible across platforms there was some discrepancy in protein sizing. This study demonstrates that protein profiling using microfluidic capillary electrophoresis is able to rapidly and reproducibly identify and characterize Yersinia species. Results show this technique is a powerful front-line, rapid and broad range screening method capable of identifying and differentiating biological agents, hoax agents and environmental bacterial species.
鼠疫耶尔森菌是一种对国家安全具有高风险的生物制剂，因其易于在人群中传播。如果鼠疫耶尔森菌被用于蓄意引发的疾病爆发，快速准确地鉴定该病原体至关重要。目前鼠疫耶尔森菌属的鉴定方法受到其对培养的依赖、获得结果所需时间和/或不适用于现场使用的方案的限制。迫切需要更快的鉴定方法。微流控毛细管电泳用于根据蛋白质谱鉴定七种鼠疫耶尔森菌属。进一步的目标包括确定在奶制品和大肠杆菌混合物中是否能检测到鼠疫耶尔森菌属，确定在盲法鉴定中是否能检测到鼠疫耶尔森菌，以及在两个平台上的重现性。对于鼠疫耶尔森菌属，在50千道尔顿（kDa）以及50至75 kDa之间检测到两条特征性蛋白带。各个鼠疫耶尔森菌属可以相互区分，并与大肠杆菌、炭疽芽孢杆菌斯特恩菌株和敌百虫（含有苏云金芽孢杆菌）区分开来。由于奶制品的蛋白质含量高，鼠疫耶尔森菌与奶制品混合时无法检测到，但与大肠杆菌混合时可以检测到。在使用12名个体的盲法程序中，物种鉴定成功率为96%。虽然蛋白质谱模式在不同平台上具有重现性，但蛋白质大小存在一些差异。本研究表明，使用微流控毛细管电泳进行蛋白质谱分析能够快速且可重复地鉴定和表征鼠疫耶尔森菌属。结果表明，该技术是一种强大的一线快速广谱筛选方法，能够鉴定和区分生物制剂、恶作剧制剂和环境细菌物种。
