M1 enrichment facilitates virus detection in patients with allogeneic hematopoietic stemcell transplantation

Abstract

Due to the severe consequences of viral infection in allogeneic hematopoietic stem cell transplantation (allo-HSCT) patients, the routine use of polymerase chain reaction (PCR) screening of viruses is common in clinic, while the sensitivities of molecular methods are not always sufficient for blood samples. We aimed to increase the detection efficiency of viremia in allo-HSCT patients with M1 bead enrichment. Blood samples of allo-HSCT patients with fever were collected. Simultaneously with analyses of real-time PCR without enrichment, M1 bead enrichment followed by real-time PCR was applied to detect possible viruses in these samples, and metagenomic next-generation sequencing analyses were also applied in 10 samples negative with real-time PCR without enrichment. Various species of viruses were detected with M1 enrichment method. Significantly, some viruses that had not been found by real-time PCR without M1 enrichment were also detected by those with M1 enrichment and verified by metagenomic next-generation sequencing analyses. Furthermore, blood samples enriched with M1 beads had lower Ct values of real-time PCR assay than those that had not been treated. In conclusion, M1 bead enrichment increased the detection efficiency of most viruses in hematological malignancy patients.

Figure 1.

M1 bead enrichment increased virus detection sensitivity in patient blood samples more efficiently than in PBS-simulated samples. (A) Real-time PCR results of PBS-simulated EBV samples analyzed with standard qPCR, M1-qPCR, or qPCR after anti-EBV antibody beads enrichment. Results are the mean standard deviation (SD) values from assays of 5 repetitions. *P < .05; (B) Real-time PCR results of 11 EBV-positive patient blood samples assayed with standard qPCR, M1-qPCR, or qPCR after anti-EBV antibody beads enrichment. (C) Diagram of M1 enrichment facilitating pathogen detection by capturing either the pathogen itself or infected leukocytes. EBV = Epstein–Barr virus, PBS = phosphate-buffered saline, PCR = polymerase chain reaction, qPCR = quantitative real-time polymerase chain reaction.