Diagnostic value of pathogen metagenomic next-generation sequencing (mNGS) for early bloodstream infections in pediatric allogeneic hematopoietic stem cell transplantation recipients

Abstract Background: Early bloodstream infection (EBSI) remains a principal determinant of morbidity and mortality after paediatric allogeneic haematopoietic stem-cell transplantation (allo-HSCT). Metagenomic next-generation sequencing (mNGS) offers a culture-independent, high-throughput approach for comprehensive pathogen identification. We investigated the diagnostic performance of plasma mNGS for EBSI in children undergoing allo-HSCT. Methods: In this single-centre, retrospective cohort study, we consecutively enrolled 161 transplant episodes performed in 157 paediatric patients at the Pediatric Hematology and Oncology Center, Institute of Hematology, Chinese Academy of Medical Sciences, between January 2024 and April 2025. Blood cultures and plasma mNGS were systematically collected from conditioning initiation to day +30 post-transplant. Group comparisons employed Fisher's exact test. Results: Blood-culture-confirmed BSI was documented in 18 of 161 episodes (11.2%). Disease-specific incidence rates were: ALL 19.4 % (6/31), AML 10.5 % (4/38), acquired aplastic anaemia/other marrow failure 7.4 % (4/54), myelodysplastic syndrome 4.5 % (1/22) and other haematologic malignancies 25.0 % (3/12). Donor-specific BSI rates were 11.5 % (7/61) after haploidentical, 15.3 % (11/72) after unrelated cord-blood and 0 % (0/34) after matched-sibling transplantation. There were no statistically significant differences were found across disease subtypes (P = 0.31) or donor sources (P = 0.60).Predominant blood-culture isolates were Streptococcus mitis, Klebsiella pneumoniae and Escherichia coli. Plasma mNGS was obtained in 39 episodes, yielding 22 positives (56.4 %). Among febrile episodes, blood culture was positive in 9/39 (23.1 %). mNGS identified bacterial infection in 8, viral infection in 6 and combined bacterial–viral infection in 8 cases. Leading bacteria were K. pneumoniae, Pseudomonas aeruginosa and S. mitis. Dominant viruses were HHV-6B, CMV and BK virus. Disease-stratified mNGS positivity was 100 % (5/5) for ALL, 75.0 % (6/8) for AML, 33.3 % (1/3) for MDS and 37.5 % (6/16) for severe aplastic anaemia. AML/ALL group exhibited significantly higher positivity than the combined MDS/SAA group (84.6 % vs 36.8 %, P = 0.017). Donor-stratified mNGS positivity was 52.1 % (12/23) after unrelated cord blood, 69.2 % (9/13) after haploidentical and 33.3 % (1/3) after matched-sibling transplantation (P > 0.05). Eight mNGS-positive patients with neurological symptoms underwent cerebrospinal fluid mNGS; seven were positive, revealing S. mitis with HHV-6 (two), HHV-6 alone (two), S. mitis alone (one), P. aeruginosa (one) and K. pneumoniae (one). Two infection-related deaths occurred before day +30. Conclusions: mNGS demonstrates high diagnostic value for early infections post-HSCT, particularly in AML/ALL patients, with superior detection of mixed and viral infections compared to blood culture. CNS involvement was common in mNGS-positive cases, highlighting the need for comprehensive pathogen screening in high-risk pediatric HSCT recipients.