Deep mining of healthy blood metagenomics and phageomes

Objective To explore the presence and potential interactions of microbes and bacteriophages in the blood of healthy individuals by employing in-depth bioinformatics mining to analyze the structure and function of the blood microbiome ecosystem. Methods Blood plasma samples from 1 600 voluntary blood donors collected at Mianyang Central Blood Station from 2012 to 2018 were subjected to DNA extraction and library construction. High-throughput sequencing was conducted using the Illumina HiSeq 4500 platform, followed by extensive bioinformatics analysis. Microbial abundance in blood samples was analyzed using metagenomic analysis software such as Bowtie2, Trimmomatic and Kraken. Subsequent phageome analysis included sequence quality control, assembly, identification, clustering and functional annotation using software such as Megahit, geNomad, CheckV and eggNOG-mapper. Phylogenetic trees, species annotation and host analysis and prediction for the identified blood bacteriophages were constructed using iTOL, BLAST and PhaBOX software. Results Metagenomic sequencing identified microbes across 36 phyla, 151 orders, 338 families, 338 genera and 3 757 species in the plasma samples. At the species level, the most abundant species included Bacillus cereus, Lactobacillus murinus, L. johnsonii, Faecalibacterium prausnitzii, B. thuringiensis, L. reuteri, Cutibacterium acnes, Dietzia sp. JS16-p6b, Mycoplasma hyorhinis, M. hyopneumoniae and Staphylococcus aureus. Through phageome analysis, 202 viral Operational Taxonomic Units (vOTUs) were identified, revealing 24 types of bacteriophages. Host analysis using the viral host database completed matches for 15 potential bacteriophage hosts, including Stenotrophomonas maltophilia, Rhodoferax lacus, Pseudoalteromonas marina, Thalassotalea loyana, Vibrio alginolyticus, V. tasmaniensis, V. vulnificus, Pseudomonas sp., Agrobacterium sp. ST15.13/040, Enterococcus gallinarum, Flavobacterium sp., Thermotoga naphthophila, Chryseobacterium sp. RU33C, L. acidipiscis and Neisseria mucosa. Conclusion The study of the healthy human blood microbiome and phageome reveals the presence of microbes and phages in the blood, which may have profound impacts on human health.