血小板制品中共存细菌全基因组测序分析及其生物学特征研究OA

Objective To establish a rapid,accurate,and decentralized workflow for bacterial whole-genome sequen-cing(WGS)and risk profiling within the shelf-life of platelet concentrates,and to characterize the species,virulence,anti-microbial resistance(AMR),and immune evasion mechanisms of co-existing bacteria in qualified platelet products,thereby providing a scientific basis for transfusion safety assessment.Methods Three units of platelet concentrates,which tested negative by routine bacterial screening,were collected from the Chengdu Blood Center between May and June 2025.Sam-ples were enriched at 37℃under six aerobic and nine anaerobic conditions for 7 days.Using a culturomics strategy,ali-quots were plated for isolation on days 1,3,5,and 7 to obtain cultivable isolates,with negative culture controls included to exclude contamination.High-molecular-weight genomic DNA was extracted via mechanical grinding,purified,and size-se-lected.Sequencing libraries were constructed and sequenced on the G-seq500 single-molecule nanopore sequencing plat-form.Genomes were assembled using Flye and polished with NextPolish,with quality evaluated by BUSCO and CheckM.Taxonomic identification was performed using GTDB-Tk.Functional annotation and database comparisons were conducted to analyze virulence factors,AMR genes,and genes related to immune evasion and environmental adaptation.Results Viable bacteria were successfully isolated from all three qualified platelet units within their shelf life.The isolates were identified as Bacillus albus,Niallia taxi,and Staphylococcus warneri.Nanopore sequencing generated 92 227-109 813 reads(totaling 680-758 Mb)with an N50 of 7 625-8 584 bp and Q20/Q30 scores of 97%/93%,respectively.All three genomes were as-sembled into complete circular chromosomes with 1-3 plasmids,achieving>93%completeness.Functional analysis re-vealed that B.albus carried multiple hemolysins,metalloproteases,and multidrug resistance genes,indicating the highest potential pathogenicity and AMR risk.S.warneri exhibited a typical multidrug resistance profile and regulatory network characteristic of coagulase-negative staphylococci,suggesting intermediate virulence.N.taxi harbored few canonical viru-lence factors and lacked functional AMR determinants,presenting a"low-virulence,low-resistance"profile.Notably,all three strains were enriched in genes encoding antimicrobial peptide resistance systems(e.g.,dltABCD,mprF,GraRS,BceAB)and antioxidant enzymes,suggesting a strong capacity to withstand immune stress in the blood environment.Con-clusion Viable bacteria can be recovered from qualified platelet concentrates that test negative by routine screening.Nano-pore WGS enables rapid strain-level identification and comprehensive risk profiling of virulence,resistance,and immune ad-aptation traits.The functional repertoires of these"co-existing"isolates range from environmental adaptation to potential pathogenicity,representing an underappreciated risk for transfusion-transmitted infections in susceptible recipients.