ABSTRACT
Eight Faecalibacterium sp. strains were isolated from feces of healthy human volunteers. Here, we describe their genome sequences. The genome sizes ranged from 2.78 Mbp to 3.23 Mbp, with an average GC content of 56.6% and encoding 2,795 protein-coding genes on average.
ANNOUNCEMENT
Faecalibacterium sp. are commensal microorganisms found ubiquitously in the human gastrointestinal tract (GIT). These microbes are important species contributing to human health through the production of butyrate, which is thought to have health-promoting properties. A reduction in Faecalibacterium in patients with different forms of inflammatory bowel disease has led researchers to believe these microorganisms confer health benefits (1–7).
This study isolated and sequenced eight strains of Faecalibacterium from human fecal samples collected in Palmerston North, New Zealand. Donors were recruited according to Massey University Ethics Approval (SOA 19/03). Volunteers were deemed healthy if they had a body mass index between 18.5 and 30; had no history of antibiotics, laxatives, or GIT infections or disorders 3 months prior to sample collection; and had moderate fiber consumption (>15 g/day). Samples were collected and processed as described by Fitzgerald et al. (8) using yeast casitone fatty acid medium supplied with glucose (YCFAG). Strain HTF-F (9) was also sequenced for comparison as a strain of interest due to its unique extrapolymeric matrix (2).
To isolate DNA, bacteria were cultured in YCFAG at 37°C overnight in an anaerobic workstation (75% N2, 15% CO2, and 5% H2; DonWhitley Scientific, UK). Samples were concentrated via centrifugation at 8,000 × g and processed using the Nucleospin soil genomic DNA purification kit (Macherey-Nagel) as per the manufacturer’s protocol. Library preparation and sequencing, including quality control (QC), was handled by Massey Genome Service (MGS; Massey University, New Zealand), using the Illumina Nextera XT kit on the Illumina MiSeq 2 × 300-bp paired-end (PE) v3 platform. Each sequence was trimmed to their longest contiguous segment within a quality cutoff (0.01), using the dynamictrim application from the SolexaQA++ software (v3.1.7.2; http://solexaqa.sourceforge.net/). Quality checking was conducted using standard parameters with FastQC (v0.11.9) (10).
For long-read sequencing, bacteria were grown again as described above, and DNA was extracted using Qiagen Genomic-tip 100/G columns per the manufacturer’s protocol. Mutanolysin (100 U; Sigma-Aldrich) and MetaPolyzyme (10 μL/sample; Sigma-Aldrich) were added to enhance bacterial lysis. Samples were sent to MGS for sample quality assessment and to Novogene (Singapore) for PacBio sequencing.
PacBio sequencing, including library preparation and QC, were performed by Novogene. The PacBio SMRTbell library was created by shearing template DNA, and the hairpin-legated dimers were purified by magnetic beads with 10-kilonucleotide size selection conditions. The library was checked with Qubit and Bioanalyzer for quantification and size distribution, respectively. Quantified libraries were pooled and sequenced on PacBio Sequel II/IIe system. The PacBio subreads and N50 values are listed in Table 1.
TABLE 1.
List of Faecalibacterium sp. strain information from this study
| Strain | Illumina read count (paired) | PacBio reads N50 (bp) | No. of PacBio subreads | GC content (%) | No. of DNA CDSa | No. of rRNAs | No. of tRNAs | Coding ratio (%) | Length (bp) | Accession no. of: |
|||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| BioSample | Genomes | Assembly | SRA | ||||||||||
| IP-1-18 | 543,181 | 14,826 | 101,178 | 56.2 | 2,814 | 18 | 64 | 86.2 | 3,038,545 | SAMN26934697 | NZ_CP094472.1 | GCA_023347355.1 | SRX15120859, SRX15120845 |
| IP-3-29 | 667,687 | 12,497 | 387,776 | 56.9 | 2,748 | 18 | 65 | 86.7 | 3,002,063 | SAMN26934698 | NZ_CP094471.1 | GCA_023347335.1 | SRX15120860, SRX15120846 |
| I2-3-92 | 704,052 | 13,398 | 279,899 | 56.8 | 2,774 | 18 | 68 | 87.1 | 2,963,404 | SAMN26934699 | NZ_CP094470.1 | GCA_023347315.1 | SRX15120853, SRX15120847 |
| HTF-F | 794,651 | 14,932 | 160,344 | 56.6 | 2,571 | 18 | 65 | 85.7 | 2,776,287 | SAMN26934700 | NZ_CP094473.1 | GCA_023347535.1 | SRX15120854, SRX15120848 |
| I3-3-33 | 596,779 | 14,126 | 491,692 | 56.6 | 2,669 | 18 | 65 | 85.4 | 2,994,777 | SAMN26934701 | NZ_CP094469.1 | GCA_023347295.1 | SRX15120855, SRX15120849 |
| I3-3-89 | 586,081 | 10,975 | 300,107 | 58 | 2,698 | 18 | 65 | 86 | 2,816,187 | SAMN26934702 | NZ_CP094468.1 | GCA_023347275.1 | SRX15120856, SRX15120850 |
| I4-1-79 | 650,123 | 15,580 | 195,760 | 56 | 3,101 | 18 | 67 | 85.7 | 3,227,950 | SAMN26934703 | NZ_CP094467.1 | GCA_023347235.1 | SRX15120857, SRX15120851 |
| I4-3-84 | 576,265 | 15,227 | 367,152 | 55.5 | 2,984 | 18 | 70 | 86.2 | 3,119,411 | SAMN26934704 | NZ_CP094466.1 | GCA_023347255.1 | SRX15120858, SRX15120852 |
CDS, coding DNA sequences.
Raw PacBio reads were filtered via Filtlong (https://github.com/rrwick/Filtlong) using a minimum subread length of 1,000 bases and a 95% cutoff. High-coverage long reads were assembled using Trycycler v0.5.3 (11), Miniasm v0.3-r179 (12), and Flye v2.9-b1768 (13) and polished with Polypolish v0.5.0 (14). Strains with low-coverage long-read data were combined with their Illumina data and hybrid assembled using Unicycler (v0.5) (12). Assembly integrity was assessed (QUAST and CheckM) on the online platform Kbase (https://kbase.us). Default parameters for all software were used. Genome annotation was performed using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP) (Table 1) (15–17). Trycycler and Unicycler confirmed all genomes to be circular.
Data availability.
All the annotated genomes and the respective long and short raw reads have been deposited in GenBank under BioProject PRJNA819544. Assembly, BioSample, and SRA details are specified in Table 1.
ACKNOWLEDGMENTS
We thank Moreno Zolfo for his helpful advice and expertise during this work and H. J. M. Harmsen for the HTF-F strain donation. D.F. was supported by a Ph.D. Fellowship from the Riddet Institute through funding provided by the New Zealand Tertiary Education Commission. The research was funded by the Ministry of Business, Innovation and Employment.
Contributor Information
Warren C. McNabb, Email: W.Mcnabb@massey.ac.nz.
Julie C. Dunning Hotopp, University of Maryland School of Medicine
REFERENCES
- 1.Miquel S, Martín R, Lashermes A, Gillet M, Meleine M, Gelot A, Eschalier A, Ardid D, Bermúdez-Humarán LG, Sokol H, Thomas M, Theodorou V, Langella PM, Carvalho FA. 2016. Anti-nociceptive effect of Faecalibacterium prausnitzii in non-inflammatory IBS-like models. Sci Rep 6:19399. doi: 10.1038/srep19399. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Rossi O, Khan MT, Schwarzer M, Hudcovic T, Srutkova D, Duncan SH, Stolte EH, Kozakova H, Flint HJ, Samsom JN, Harmsen HJM, Wells JM. 2015. Faecalibacterium prausnitzii strain HTF-F and its extracellular polymeric matrix attenuate clinical parameters in DSS-induced colitis. PLoS One 10:e0123013. doi: 10.1371/journal.pone.0123013. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Laval L, Martin R, Natividad JNF, Chain FC, Miquel S, Desclée de Maredsous C, Capronnier S, Sokol H, Verdu EF, van Hylckama Vlieg JET, Bermúdez-Humáran LG, Smokvina T, Langella P, Chain F, Miquel S, Desclée de Maredsous C, Capronnier S, Sokol H, VerdU EF, van Hylckama Vlieg JET, Bermúdez-Humarán LG, Smokvina T, Langella P. 2015. Lactobacillus rhamnosus CNCM I-3690 and the commensal bacterium faecalibacterium prausnitzii A2-165 exhibit similar protective effects to induced barrier hyper-permeability in mice. Gut Microbes 6:1–9. doi: 10.4161/19490976.2014.990784. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Martín R, Miquel S, Chain F, Natividad JM, Jury J, Lu J, Sokol H, Theodorou V, Bercik P, Verdu EF, Langella P, Bermúdez-Humarán LG. 2015. Faecalibacterium prausnitzii prevents physiological damages in a chronic low-grade inflammation murine model. BMC Microbiol 15:67. doi: 10.1186/s12866-015-0400-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Quévrain E, Maubert MA, Michon C, Chain F, Marquant R, Tailhades J, Miquel S, Carlier L, Bermúdez-Humarán LG, Pigneur B, Lequin O, Kharrat P, Thomas G, Rainteau D, Aubry C, Breyner N, Afonso C, Lavielle S, Grill J-P, Chassaing G, Chatel JM, Trugnan G, Xavier R, Langella PM, Sokol H, Seksik P. 2016. Identification of an anti-inflammatory protein from Faecalibacterium prausnitzii, a commensal bacterium deficient in Crohn’s disease. Gut 65:415–425. doi: 10.1136/gutjnl-2014-307649. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Bridonneau C, Altare F, Tabiasco J, Alameddine J, Godefroy E, Sokol H, Yazdanbakhsh K, Papargyris L, Sarrabayrouse G, Jotereau F. 2019. Faecalibacterium prausnitzii Skews Human DC to Prime IL10-Producing T Cells Through TLR2/6/JNK Signaling and IL-10, IL-27, CD39, and IDO-1 Induction. Front Immunol 10:143. doi: 10.3389/fimmu.2019.00143. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Hu W, Lu W, Li L, Zhang H, Lee Y, Chen W, Zhao J. 2021. Both living and dead Faecalibacterium prausnitzii alleviate house dust mite-induced allergic asthma through the modulation of gut microbiota and short-chain fatty acid production. J Sci Food Agric 101:5563–5573. doi: 10.1002/jsfa.11207. [DOI] [PubMed] [Google Scholar]
- 8.Fitzgerald CB, Shkoporov AN, Sutton TDS, Chaplin AV, Velayudhan V, Ross RP, Hill C. 2018. Comparative analysis of Faecalibacterium prausnitzii genomes shows a high level of genome plasticity and warrants separation into new species-level taxa. BMC Genomics 19:931. doi: 10.1186/s12864-018-5313-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9.Lopez-Siles M, Khan TM, Duncan SH, Harmsen HJM, Garcia-Gil LJ, Flint HJ. 2012. Cultured representatives of two major phylogroups of human colonic Faecalibacterium prausnitzii can utilize pectin, uronic acids, and host-derived substrates for growth. Appl Environ Microbiol 78:420–428. doi: 10.1128/AEM.06858-11. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Andrews S. 2010. FastQC: a quality control tool for high throughput sequence data. https://www.bioinformatics.babraham.ac.uk/projects/fastqc/.
- 11.Wick RR, Judd LM, Cerdeira LT, Hawkey J, Méric G, Vezina B, Wyres KL, Holt KE. 2021. Trycycler: consensus long-read assemblies for bacterial genomes. Genome Biol 22:266. doi: 10.1186/s13059-021-02483-z. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Wick RR, Judd LM, Gorrie CL, Holt KE. 2017. Unicycler: resolving bacterial genome assemblies from short and long sequencing reads. PLoS Comput Biol 13:e1005595. doi: 10.1371/journal.pcbi.1005595. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Kolmogorov M, Yuan J, Lin Y, Pevzner PA. 2019. Assembly of long, error-prone reads using repeat graphs. Nat Biotechnol 37:540–546. doi: 10.1038/s41587-019-0072-8. [DOI] [PubMed] [Google Scholar]
- 14.Wick RR, Holt KE. 2022. Polypolish: short-read polishing of long-read bacterial genome assemblies. PLoS Comput Biol 18:e1009802. doi: 10.1371/journal.pcbi.1009802. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15.Haft DH, DiCuccio M, Badretdin A, Brover V, Chetvernin V, O'Neill K, Li W, Chitsaz F, Derbyshire MK, Gonzales NR, Gwadz M, Lu F, Marchler GH, Song JS, Thanki N, Yamashita RA, Zheng C, Thibaud-Nissen F, Geer LY, Marchler-Bauer A, Pruitt KD. 2018. RefSeq: an update on prokaryotic genome annotation and curation. Nucleic Acids Res 46:D851–D860. doi: 10.1093/nar/gkx1068. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Li W, O'Neill KR, Haft DH, DiCuccio M, Chetvernin V, Badretdin A, Coulouris G, Chitsaz F, Derbyshire MK, Durkin AS, Gonzales NR, Gwadz M, Lanczycki CJ, Song JS, Thanki N, Wang J, Yamashita RA, Yang M, Zheng C, Marchler-Bauer A, Thibaud-Nissen F. 2021. RefSeq: expanding the Prokaryotic Genome Annotation Pipeline reach with protein family model curation. Nucleic Acids Res 49:D1020–D1028. doi: 10.1093/nar/gkaa1105. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 17.Tatusova T, Dicuccio M, Badretdin A, Chetvernin V, Nawrocki EP, Zaslavsky L, Lomsadze A, Pruitt KD, Borodovsky M, Ostell J. 2016. NCBI Prokaryotic Genome Annotation Pipeline. Nucleic Acids Res 44:6614–6624. doi: 10.1093/nar/gkw569. [DOI] [PMC free article] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
All the annotated genomes and the respective long and short raw reads have been deposited in GenBank under BioProject PRJNA819544. Assembly, BioSample, and SRA details are specified in Table 1.