Complete genome sequence of Peptostreptococcus anaerobius SB204, isolated from a human colonic adenocarcinoma

TL;DR

This paper presents the full genome sequence of a Peptostreptococcus anaerobius strain found in a colorectal cancer tumor.

Contribution

The complete genome sequence of Peptostreptococcus anaerobius SB204 is newly reported.

Findings

The genome is a single chromosomal contig of 2.15 Mbp.

The GC content of the genome is 36.1%.

Abstract

We report the complete genome sequence of Peptostreptococcus anaerobius SB204, a strain isolated from the resected tumor of a treatment-naive patient with colorectal cancer. The genome comprises a single chromosomal contig of 2.15 Mbp with an overall GC content of 36.1%.

Figures

Heat map of ANI values among selected P. anaerobius genomes. (Left) Reference-free whole-genome phylogenetic tree of P. anaerobius genomes (n = 22) generated with kSNP4 (11) (kmer = 17, FCK = 0.658) with (right) corresponding heatmap of ANI values. Duplicate genomes (ANI > 99%) have been removed. ANI heatmap was generated with the pheatmap R package, RStudio version 2024.12.0+467. The tree was visualized using iTOL (12), and the figure was made with Biorender.

Tables

TABLE 1. Publicly available genome assemblies used for ANI analysis

Strain	Accession no.	Isolation source
CD12_MAG18	GCA_019013055.1	Mucosal tissue from Crohn’s disease patient
SRR11749280_bin.18_metaWRAP_v1.3_MAG	GCA_947086575.1	Vaginal metagenome
UMB10161C	GCA_030227555.1	Urine
SRR16916875_bin.10_metaWRAP_v1.3_MAG	GCF_946998135.1	Vaginal metagenome
PA 653-L	GCF_000178095.1	Biological product
N4_154_020G1_dasN4_154_020G1_abawaca.36	GCA_032475295.1	Infant feces
2225st2_D9_2225SCRN_200828	GCA_039756995.1	NAa
RTP21358st1_C2_RTP21358_211008	GCA_040909105.1	NA
DFI.6.106	GCA_024462755.1	Fecal sample
CC00978	GCA_964251445.1	NA
SRR17635664_bin.8_metaWRAP_v1.3_MAG	GCA_947253115.1	NA
MJR8628A	GCA_001563595.1	Vagina
N3_174_000G1_dasN3_174_000G1_concoct_28	GCA_032552295.1	Infant feces
MGS621	GCA_000431415.1	NA
SRR3546782_bin.52_CONCOCT_v1.1_MAG	GCA_937974095.1	NA
E6_m1001271B151109d1_201121	GCA_028326525.1	NA
N2_065_000G1_dasN2_065_000G1_abawaca.31	GCA_032557275.1	Infant feces
N4_179_023G1_dasN4_179_023G1_concoct_1	GCA_032472735.1	Infant feces
KA00810	GCA_001553145.1	Vagina
BSD2780120874b_170522_E5	GCA_015548685.1	NA
N4_205_052G1_dasN4_205_052G1_concoct_1	GCA_032471295.1	Infant feces

Full text

ANNOUNCEMENT

Peptostreptococcus anaerobius was originally classified under Streptococcus in 1905 but reclassified to Peptostreptococcus in 1936 due to its distinct anaerobic growth and metabolic characteristics (1). It is a gram-positive, anaerobic, non-spore-forming coccus commonly found in the human gastrointestinal and genitourinary tracts (2–4). Although typically regarded as a commensal organism, it is implicated in a range of infections, including pelvic inflammatory disease (5), dental abscesses (6), intra-abdominal infections (7), and bacteremia (8). Here, we document the isolation and complete genome sequence of P. anaerobius strain SB204 from colorectal adenocarcinoma tissue of a treatment-naive female patient.

With written informed patient consent, approved by the Fred Hutchinson Cancer Center Institutional Review Board (protocol RG-1006552), SB204 was isolated from resected tumor specimen plated on Fastidious Anaerobic Agar (FAA; Oxoid, Thermo Fisher Scientific) supplemented with 10% defibrinated horse blood (Lampire Biological Laboratories, Fisher Scientific) and incubated under anaerobic conditions at 37°C for 48 h (AnaeroGen Gas Generating Systems, Oxoid, Thermo Fisher Scientific). BLAST analysis of the SB204 16S rRNA gene sequence determined initial taxonomic classification as P. anaerobius. High-molecular-weight genomic DNA was extracted using the MasterPure DNA Purification Kit (Epicentre, Lucigen). DNA concentration and quality were measured using the Qubit dsDNA BR Assay Kit (Thermo Fisher Scientific). DNA was sheared via G-tube (Covaris) as previously described (9) and size selected for fragments >5 kb using BluePippin (Sage Science). Sequencing library was prepared using the HiFi Assembly Kit (Pacific Biosciences). Single-molecule real-time (SMRT) sequencing (10) with base kinetics was performed on a PacBio Sequel I instrument with V3 sequencing chemistry (Pacific Biosciences). Sequencing reads were quality filtered and processed using microbial assembler in the Pacific Biosciences’ SMRTAnalysis pipeline (version 9.0.0.92188) with default parameters. Genome assembly was performed using Microbial Assembler, which included error correction and chromosomal rotation to place dnaA at the origin. Genome assembly showed 93,738 polymerase reads that were further partitioned into 591,799 mapped subreads, with an N50 value of 9,175 nucleotides and a total of 4,890,837,089 subread bases, yielding a mean coverage of 2,212×. Average nucleotide identity (ANI) analysis against publicly available P. anaerobius genomes (Table 1, Fig. 1) supported its species-level identification.

Heat map of ANI values among selected P. anaerobius genomes. (Left) Reference-free whole-genome phylogenetic tree of P. anaerobius genomes (n = 22) generated with kSNP4 (11) (kmer = 17, FCK = 0.658) with (right) corresponding heatmap of ANI values. Duplicate genomes (ANI > 99%) have been removed. ANI heatmap was generated with the pheatmap R package, RStudio version 2024.12.0+467. The tree was visualized using iTOL (12), and the figure was made with Biorender.

Genome annotation using the NCBI Prokaryotic Genome Annotation Pipeline (13) (v6.1) identified 1,872 coding sequences, a GC content of 36.1%, and 78 RNAs. PADLOC (14) (v2.0.0) and CRISPR-Cas (15) analysis predicted the presence of multiple defense systems, including Type I-B CRISPR-Cas and Type I restriction-modification (RM) systems. To identify putative nucleotide motifs modified by the Type I RM system, we performed methylome analysis via REBASE (16), which identified m6A modification of TAAYN_5_CTC/CAGN_5_RRT. Analysis via the Comprehensive Antibiotic Resistance Database (17) (v4.0.1) identified antimicrobial resistance genes associated with putative resistance to fluoroquinolones (sdrM), glycopeptides (vanT and vanW), and tetracycline (tetM).

Currently, there are 51 publicly available P. anaerobius genome assemblies, although by ANI analysis, some appear to be duplicates. Notably, SB204 is the first complete genome sequence available for this species and the first reported strain from colorectal tumor tissue (Table 1). This isolate and its genome provide a novel resource for studying the clinical role(s) of this organism, particularly in the tumor microenvironment.