Abstract：
UNASSIGNED: Recombinase-aided polymerase chain reaction (RAP) is a sensitive, single-tube, two-stage nucleic acid amplification method. This study aimed to develop an assay that can be used for the early diagnosis of three types of bacteremia caused by Staphylococcus aureus (SA), Pseudomonas aeruginosa (PA), and Acinetobacter baumannii (AB) in the bloodstream based on recombinant human mannan-binding lectin protein (M1 protein)-conjugated magnetic bead (M1 bead) enrichment of pathogens combined with RAP.
UNASSIGNED: Recombinant plasmids were used to evaluate the assay sensitivity. Common blood influenza bacteria were used for the specific detection. Simulated and clinical plasma samples were enriched with M1 beads and then subjected to multiple recombinase-aided PCR (M-RAP) and quantitative PCR (qPCR) assays. Kappa analysis was used to evaluate the consistency between the two assays.
UNASSIGNED: The M-RAP method had sensitivity rates of 1, 10, and 1 copies/μL for the detection of SA, PA, and AB plasmids, respectively, without cross-reaction to other bacterial species. The M-RAP assay obtained results for < 10 CFU/mL pathogens in the blood within 4 h, with higher sensitivity than qPCR. M-RAP and qPCR for SA, PA, and AB yielded Kappa values of 0.839, 0.815, and 0.856, respectively ( P < 0.05).
UNASSIGNED: An M-RAP assay for SA, PA, and AB in blood samples utilizing M1 bead enrichment has been developed and can be potentially used for the early detection of bacteremia.
UNASSIGNED: Recombinant plasmids were used to evaluate the assay sensitivity. Common blood influenza bacteria were used for the specific detection. Simulated and clinical plasma samples were enriched with M1 beads and then subjected to multiple recombinase-aided PCR (M-RAP) and quantitative PCR (qPCR) assays. Kappa analysis was used to evaluate the consistency between the two assays.
UNASSIGNED: The M-RAP method had sensitivity rates of 1, 10, and 1 copies/μL for the detection of SA, PA, and AB plasmids, respectively, without cross-reaction to other bacterial species. The M-RAP assay obtained results for < 10 CFU/mL pathogens in the blood within 4 h, with higher sensitivity than qPCR. M-RAP and qPCR for SA, PA, and AB yielded Kappa values of 0.839, 0.815, and 0.856, respectively ( P < 0.05).
UNASSIGNED: An M-RAP assay for SA, PA, and AB in blood samples utilizing M1 bead enrichment has been developed and can be potentially used for the early detection of bacteremia.
摘要：
■重组酶辅助聚合酶链反应（RAP）是一种敏感的，单管,二阶段核酸扩增法。本研究旨在开发一种可用于早期诊断由金黄色葡萄球菌（SA）引起的三种类型的菌血症的方法。铜绿假单胞菌(PA),基于重组人甘露聚糖结合凝集素蛋白(M1蛋白)结合磁珠(M1微珠)富集病原菌并结合RAP。
■重组质粒用于评估测定灵敏度。采用普通血液流感细菌进行特异性检测。用M1珠富集模拟和临床血浆样品,然后进行多重重组酶辅助PCR(M-RAP)和定量PCR(qPCR)测定。使用Kappa分析来评估两种测定之间的一致性。
■M-RAP方法检测SA的灵敏度为1、10和1拷贝/μL，PA,和AB质粒，分别,与其他细菌物种没有交叉反应。M-RAP分析在4小时内获得血液中<10CFU/mL病原体的结果，具有比qPCR更高的灵敏度。SA的M-RAP和qPCR，PA,和AB的Kappa值分别为0.839、0.815和0.856（P<0.05）。
■SA的M-RAP测定，PA,已经开发了利用M1珠富集的血液样品中的AB，并且可以潜在地用于菌血症的早期检测。
■重组质粒用于评估测定灵敏度。采用普通血液流感细菌进行特异性检测。用M1珠富集模拟和临床血浆样品,然后进行多重重组酶辅助PCR(M-RAP)和定量PCR(qPCR)测定。使用Kappa分析来评估两种测定之间的一致性。
■M-RAP方法检测SA的灵敏度为1、10和1拷贝/μL，PA,和AB质粒，分别,与其他细菌物种没有交叉反应。M-RAP分析在4小时内获得血液中<10CFU/mL病原体的结果，具有比qPCR更高的灵敏度。SA的M-RAP和qPCR，PA,和AB的Kappa值分别为0.839、0.815和0.856（P<0.05）。
■SA的M-RAP测定，PA,已经开发了利用M1珠富集的血液样品中的AB，并且可以潜在地用于菌血症的早期检测。
