RAPID DETERMINATION OF ANTIMICROBIAL SUSCEPTIBILITY BY FLOW CYTOMETRY

(1)抗菌药物敏感性的快速测定研究了流式细胞术在加速药敏试验中的应用。首先，开发了一种新的流式细胞仪(MFCM)，用于检测细菌等微小颗粒。MFCM测量粒子计数、脉冲强度和前向散射光的宽度(分别为FSC和FSCW)。用Microsoft Excel软件分析FSC与FSCW的双参数直方图(DPH)。分析了12种细菌溶液在37℃剧烈摇动后DPH上的颗粒分布，其中存在和不存在连续稀释的反式微生物(PIPC、LVFX、AMK和Mino)。培养后，PIPC敏感微生物向FSC和FSCW较大的区域转移，而LVFX、AMK和Mino敏感细菌聚集到FSC较小的区域。通过将这些一致的观察结果与金标准微量稀释MIC进行比较，制定了在2小时内确定药物敏感性的定量标准，特别是对革兰氏阴性杆菌。(2)细菌感染的快速诊断在尿路感染中，尿液经常被细菌等微小颗粒严重污染。为了区分细菌和非细菌颗粒，尿样在37℃下剧烈摇动1-2小时，从孵育后的DPH中减去孵育前的DPH。减去DPH后的净变化表明细菌的生长。这种方法可以在微生物成倍繁殖之前对菌尿的诊断做出贡献。

(1) RAPID DETERMINATION OF ANTIMICROBIAL SUSCEPTIBILITYThe applicability of flow cytometry was studied to accelerate antimicrobial susceptibility testing. First, a new flow cytometer (MFCM) was developed for detection of as small particles as bacteria. MFCM measures the particle count, the pulse intensity and width of forward scattered light (Fsc and Fscw, respectively). Microsoft Excel software was used to analyze dual-parameter histogram (DPH) of Fsc vs. Fscw. Particle distributions on DPH were analyzed after incubation of bacterial solutions (12 species) with vigorous shaking at 37C in the presence and absence of serially diluted anatimicrobials (PIPC, LVFX, AMK and MINO). After incubation, PIPC-sensitive microbes shifted to the area with larger Fsc and Fscw, while LVFX-, AMK-and MINO-sensitive bacteria gathered to the area with smaller FSC. By comparing these consistent observations with the gold standard microdilution MICs, the quantitative criteria was developed for determining antimicrobial susceptibility within 2 hours, especially for gram-negative bacilli.(2) RAPID DIAGNOSIS OF BACTERIURIAIn urinary tract infection, urine is often heavily contaminated with particles as small as bacteria. To differentiate bacteria with non-bacterial particles, urine samples were vigorously shaked for 1-2 hours at 37C and pre-incubation DPH was subtracted from post-incubation DPH. The net changes on subtracted DPH were indicative of bacterial growth. This method could contribute to the diagnosis of bacteriuria before microbes multiplied severalfold.