An unclassified Enterobacter strain was isolated from the pitcher fluids of a Sarracenia rosea pitcher plant growing at Splinter Hill Bog in Alabama. Its genome was sequenced using the Illumina platform. A genome assembly of 4,673,815 bp was obtained. In total, 4,646 protein-encoding sequences and 72 RNA genes are predicted from this assembly.
ABSTRACT
An unclassified Enterobacter strain was isolated from the pitcher fluids of a Sarracenia rosea pitcher plant growing at Splinter Hill Bog in Alabama. Its genome was sequenced using the Illumina platform. A genome assembly of 4,673,815 bp was obtained. In total, 4,646 protein-encoding sequences and 72 RNA genes are predicted from this assembly.
ANNOUNCEMENT
The genus Enterobacter was first proposed in 1960 (1). Enterobacter belongs to the Enterobacteriaceae, a group of Gram-negative, facultatively anaerobic, motile, and straight-rod bacteria (2). Enterobacter species are widespread in aquatic environments and are also present in the intestine of animals (3). Enterobacter species are sometimes found as pathogens that cause nosocomial infections such as urinary tract infections and bacteremia (4).
The strain used in this study was isolated from the fluids of Sarracenia rosea pitcher plants from Splinter Hill Bog (Alabama) in March of 2012. Pitcher fluids were plated onto medium used for the culture of Pyrococcus furiosus (5) and streaked from single colonies onto the same medium. An individual colony was then picked into nutrient broth for growth and eventual storage at −80°C in 25% glycerol. DNA was extracted using the Quick-DNA fungal/bacterial miniprep kit (catalog number D6005; Zymo Research). A paired-end library was prepared using the Nextera XT DNA library preparation kit v2 (catalog number FC-131-1002; Illumina) with an average insert size of ∼250 bp for Illumina MiSeq sequencing. Out of 14,784,154 Illumina raw reads, we retained 4,889,590 reads after quality filtering with Trimmomatic (version 0.36) (6). FastQC (version 0.11.4) was employed to evaluate the quality of the cleaned reads (7). SOAPdenovo2 (version r240; parameter -K 83) was used to assemble the reads into contigs (8). In total, 252 contigs (all >400 bp in length), with sequence coverage of more than 20×, were obtained. Both the NCBI Prokaryotic Genome Annotation Pipeline (9) and the online RAST server (RASTtk Toolkit) were used for annotation (10). Annotation by the NCBI Prokaryotic Genome Annotation Pipeline was provided with the GenBank submission. Default parameters were used for all software tools unless otherwise noted.
The final assembly has 4,673,815 bp. The GC content is ∼55.9%. The N50 and L50 values are 41,959 bp and 33, respectively. The sequence depth of the genome assembly is ∼260×. The genome contains a predicted 4,718 genes, which consisted of 4,646 protein-coding sequences and 72 RNA genes. About 38% of the protein-coding sequences (1,764 genes) for this genome were grouped in the functional subsystems of the RAST SEED server (10). Beta-lactam resistance gene blaACT-6 and fosfomycin resistance gene fosA were predicted by the Center for Genomic Epidemiology (CGE)’s ResFinder (11). The strain was predicted to have a 75.5% probability of being a human pathogen according to CGE’s PathogenFinder (12). The two most closely related genomes, discovered by an NCBI BLASTn analysis using the nucleotide collection (nt/nr) database, are Enterobacter roggenkampii strain R11 (GenBank accession number NZ_CP019839) and Enterobacter cloacae subsp. cloacae ATCC 13047 (CP001918). The average nucleotide identities (ANI) of these two strains compared to Enterobacter sp. C6 are 93.7% and 87.9%, respectively (13). Several recent studies have demonstrated that a >95% ANI value is indicative of the same species (14). Both ANI values between Enterobacter sp. C6 and either E. cloacae strain are <95%, which suggests that Enterobacter sp. C6 is a new species of Enterobacter.
Data availability.
This whole-genome shotgun project has been deposited in GenBank under accession number QPFW00000000. The raw read accession number is SRX6867760. The version described in this paper is the first version, QPFW01000000. The BioProject number is PRJNA476218, and the BioSample number is SAMN09428499.
ACKNOWLEDGMENTS
We thank Willie L. Rogers and Russell L. Malmberg for providing pitcher fluids collected from the Splinter Hill Bog, Sanjeev Chandrayan for his help with anaerobic culturing, and Macrogen, Inc., for Illumina sequencing.
This research was supported by the Giles Endowment (to J.L.B.).
REFERENCES
- 1.Hormaeche E, Edwards P. 1960. A proposed genus Enterobacter. Int Bull Bacteriol Nomencl Taxon 10:71–74. doi: 10.1099/0096266X-10-2-71. [DOI] [Google Scholar]
- 2.Tan W-S, Yunos N, Tan P-W, Mohamad N, Adrian T-G-S, Yin W-F, Chan K-G. 2014. Freshwater-borne bacteria isolated from a Malaysian rainforest waterfall exhibiting quorum sensing properties. Sensors (Basel) 14:10527–10537. doi: 10.3390/s140610527. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Cabral JP. 2010. Water microbiology. Bacterial pathogens and water. Int J Environ Res Public Health 7:3657–3703. doi: 10.3390/ijerph7103657. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Grimont F, Grimont PA. 2006. The genus Enterobacter, p 197–214. In Dworkin M, Falkow S, Rosenberg E, Schleifer K-H, Stackebrandt E (ed), The prokaryotes, vol 6, Proteobacteria: gamma subclass. Springer, New York, NY. [Google Scholar]
- 5.Adams MW, Holden JF, Menon AL, Schut GJ, Grunden AM, Hou C, Hutchins AM, Jenney FE, Kim C, Ma K, Pan G, Roy R, Sapra R, Story SV, Verhagen MF. 2001. Key role for sulfur in peptide metabolism and in regulation of three hydrogenases in the hyperthermophilic archaeon Pyrococcus furiosus. J Bacteriol 183:716–724. doi: 10.1128/JB.183.2.716-724.2001. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Bolger AM, Lohse M, Usadel B. 2014. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30:2114–2120. doi: 10.1093/bioinformatics/btu170. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Andrews S. 2010. FastQC: a quality control tool for high throughput sequence data. Babraham Bioinformatics, Cambridge, United Kingdom. [Google Scholar]
- 8.Luo R, Liu B, Xie Y, Li Z, Huang W, Yuan J, He G, Chen Y, Pan Q, Liu Y, Tang J, Wu G, Zhang H, Shi Y, Liu Y, Yu C, Wang B, Lu Y, Han C, Cheung DW, Yiu S-M, Peng S, Xiaoqian Z, Liu G, Liao X, Li Y, Yang H, Wang J, Lam T-W, Wang J. 2012. SOAPdenovo2: an empirically improved memory-efficient short-read de novo assembler. Gigascience 1:18. doi: 10.1186/2047-217X-1-18. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9.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]
- 10.Aziz RK, Bartels D, Best AA, DeJongh M, Disz T, Edwards RA, Formsma K, Gerdes S, Glass EM, Kubal M, Meyer F, Olsen GJ, Olson R, Osterman AL, Overbeek RA, McNeil LK, Paarmann D, Paczian T, Parrello B, Pusch GD, Reich C, Stevens R, Vassieva O, Vonstein V, Wilke A, Zagnitko O. 2008. The RAST server: Rapid Annotations using Subsystems Technology. BMC Genomics 9:75. doi: 10.1186/1471-2164-9-75. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Zankari E, Hasman H, Cosentino S, Vestergaard M, Rasmussen S, Lund O, Aarestrup FM, Larsen MV. 2012. Identification of acquired antimicrobial resistance genes. J Antimicrob Chemother 67:2640–2644. doi: 10.1093/jac/dks261. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Cosentino S, Larsen MV, Aarestrup FM, Lund O. 2013. PathogenFinder—distinguishing friend from foe using bacterial whole genome sequence data. PLoS One 8:e77302. doi: 10.1371/journal.pone.0077302. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Rodriguez-R L, Konstantinidis K. 2014. Bypassing cultivation to identify bacterial species. Microbe 9:111–118. doi: 10.1128/microbe.9.111.1. [DOI] [Google Scholar]
- 14.Jain C, Rodriguez-R LM, Phillippy AM, Konstantinidis KT, Aluru S. 2018. High throughput ANI analysis of 90K prokaryotic genomes reveals clear species boundaries. Nat Commun 9:5114. doi: 10.1038/s41467-018-07641-9. [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
This whole-genome shotgun project has been deposited in GenBank under accession number QPFW00000000. The raw read accession number is SRX6867760. The version described in this paper is the first version, QPFW01000000. The BioProject number is PRJNA476218, and the BioSample number is SAMN09428499.