Great Ormond Street Institute of Child Health, London, United Kingdom.
Department of Mathematics, Imperial College London, London, United Kingdom.
PLoS One. 2022 Nov 22;17(11):e0276777. doi: 10.1371/journal.pone.0276777. eCollection 2022.
Rapid identification of potentially life-threatening blood stream infections (BSI) improves clinical outcomes, yet conventional blood culture (BC) identification methods require ~24-72 hours of liquid culture, plus 24-48 hours to generate single colonies on solid media suitable for identification by mass spectrometry (MS). Newer rapid centrifugation techniques, such as the Bruker MBT-Sepsityper® IVD, replace culturing on solid media and expedite the diagnosis of BCs but frequently demonstrate reduced sensitivity for identifying clinically significant Gram-positive bacterial or fungal infections. This study introduces a protocol that utilises the broad-range binding properties of an engineered version of mannose-binding lectin linked to the Fc portion of immunoglobulin (FcMBL) to capture and enrich pathogens combined with matrix-assisted laser desorption-ionisation time-of-flight (MALDI-TOF) MS for enhanced infection identification in BCs. The FcMBL method identified 94.1% (64 of 68) of clinical BCs processed, with a high sensitivity for both Gram-negative and Gram-positive bacteria (94.7 and 93.2%, respectively). The FcMBL method identified more patient positive BCs than the Sepsityper® (25 of 25 vs 17 of 25), notably with 100% (3/3) sensitivity for clinical candidemia, compared to only 33% (1/3) for the Sepsityper®. Additionally, during inoculation experiments, the FcMBL method demonstrated a greater sensitivity, identifying 100% (24/24) of candida to genus level and 9/24 (37.5%) top species level compared to 70.8% (17/24) to genus and 6/24 to species (25%) using the Sepsityper®. This study demonstrates that capture and enrichment of samples using magnetic FcMBL-conjugated beads is superior to rapid centrifugation methods for identification of BCs by MALDI-TOF MS. Deploying the FcMBL method therefore offers potential clinical benefits in sensitivity and reduced turnaround times for BC diagnosis compared to the standard Sepsityper® kit, especially for fungal diagnosis.
快速鉴定潜在危及生命的血流感染（BSI）可改善临床结局，但传统的血培养（BC）鉴定方法需要约 24-72 小时的液体培养，加上 24-48 小时才能在固体培养基上生成适合质谱（MS）鉴定的单个菌落。较新的快速离心技术，如 Bruker MBT-Sepsityper® IVD，替代了固体培养基培养，并加快了 BC 的诊断速度，但经常显示出降低的敏感性，无法识别临床上重要的革兰氏阳性细菌或真菌感染。本研究介绍了一种利用甘露糖结合凝集素的工程化版本与免疫球蛋白的 Fc 部分（FcMBL）结合的广谱结合特性来捕获和富集病原体，然后结合基质辅助激光解吸电离飞行时间（MALDI-TOF）MS 来增强 BC 中感染鉴定的方案。FcMBL 方法鉴定了处理的 94.1%（68 个中的 64 个）临床 BC，对革兰氏阴性和革兰氏阳性细菌均具有高敏感性（分别为 94.7%和 93.2%）。FcMBL 方法鉴定的阳性 BC 患者比 Sepsityper®（25 个中的 25 个比 17 个中的 25 个）更多，特别是对临床念珠菌血症的敏感性为 100%（3/3），而 Sepsityper®仅为 33%（1/3）。此外，在接种实验中，FcMBL 方法的敏感性更高，可鉴定出 100%（24/24）的念珠菌属水平和 9/24（37.5%）的顶级物种水平，而 Sepsityper®为 70.8%（24/24）的属水平和 6/24 的种水平（25%）。本研究表明，与快速离心方法相比，使用磁性 FcMBL 偶联珠对样品进行捕获和富集更有利于通过 MALDI-TOF MS 鉴定 BC。与标准 Sepsityper®试剂盒相比，FcMBL 方法在 BC 诊断的敏感性和周转时间方面具有潜在的临床优势，特别是在真菌诊断方面。
