Open Access

Draft genome sequence of Desulfoplanes formicivorans Pf12BT, a sulfate-reducing bacterium of the family Desulfomicrobiaceae

Standards in Genomic Sciences201712:34

https://doi.org/10.1186/s40793-017-0246-2

Received: 6 July 2016

Accepted: 26 May 2017

Published: 5 June 2017

Abstract

Desulfoplanes formicivorans strain Pf12BT is the type strain of the type species in the genus Desulfoplanes, which is the one of the genera in the family Desulfomicrobiaceae within the order Desulfovibrionales. This deltaproteobacterium was isolated from a blackish meromictic lake sediment. D. formicivorans strain Pf12BT is a Gram-negative, motile and sulfate-reducing bacterium. Cells of strain Pf12BT are characterized by possession of vibroid morphology and red fluorescent pigment. Here we describe the features, draft genome sequence and annotation of this organism, the sole species of the genus Desulfoplanes. The genome comprised 3,000,979 bp, 2,657 protein-coding genes and 58 RNA genes.

Keywords

Bacteria Gram-negative Anaerobe Sulfate-reducer Desulfomicrobiaceae

Introduction

Strain Pf12BT (= NBRC 110391 T = DSM 28890 T) is the type strain of Desulfoplanes formicivorans , which is the type species of the genus Desulfoplanes in the family Desulfomicrobiaceae . The family Desulfomicrobiaceae was proposed by Kuever et al. (2006) and contained only one genus, Desulfomicrobium . The genus Desulfoplanes was later added to this family because of the phylogenetic position [1]. All members of the family Desulfomicrobiaceae including D. formicivorans are sulfate reducers and incomplete oxidizers, which are unable to completely oxidize organic matters to CO2. All known strains of the genus Desulfomicrobium have rod- or ellipsoidal-shaped morphology and they all lack desulfoviridin, which is a red fluorescent pigment [24]. In contrast, D. formicivorans strain Pf12BT was characterized by vibroid morphology and possession of red fluorescent pigment.

In this study we summarize the features of D. formicivorans strain Pf12BT and provide an overview of the draft genome sequence and annotation of this strain.

Organism Information

Classification and features

D. formicivorans strain Pf12BT was isolated from the anaerobic sediments of a meromictic lake [1, 5]. Cells of this strain are Gram-negative, motile, non-spore-forming and vibroids (Fig 1, Table 1). Under UV illumination, cell lysate of the strain exhibited red fluorescence suggesting the presence of desulfoviridin. Temperature range for growth is 13–50 °C, with an optimum temperature at 42–45 °C. NaCl concentration for growth is 0.5–8% (w/v) and optimal concentration is 1–4% (w/v). This bacterium is strictly anaerobic and is capable of respiration and fermentation. Sulfate, thiosulfate and sulfite are used as electron acceptors for growth. Nitrate is not used for respiration. Pyruvate, malate and fumarate are used for fermentative growth.
Fig. 1

Photomicrograph of cells of D. formicivorans strain Pf12BT. Cells were grown with formate (10 mM) and yeast extract (0.5 g l-1) in the presence of sulfate for 2 days

Table 1

Classification and general features of Desulfoplaens formicivorans strain Pf12BT according to MIGS recommendations

MIGS ID

Property

Term

Evidence codea

 

Classification

Domain Bacteria

TAS [6]

  

Phylum Proteobacteria

TAS [18]

  

Class Deltaproteobacteria

TAS [19, 20]

  

Order Desulfovibrionales

TAS [20, 21]

  

Family Desulfomicrobiaceae

TAS [4, 20]

  

Genus Desulfoplanes

TAS [1]

  

Species Desulfoplanes formicivorans

TAS [1]

  

Type strain: Pf12BT (DSM 28890)

 
 

Gram stain

negative

TAS [1]

 

Cell shape

vibroid

TAS [1]

 

Motility

motile

TAS [1]

 

Sporulation

nonsporulating

TAS [1]

 

Temperature range

13–50 °C

TAS [1]

 

Optimum temperature

42–45 °C

TAS [1]

 

pH range; Optimum

6.1–8.6; 7.0–7.5

TAS [1]

 

Carbon source

organic acids

TAS [1]

MIGS-6

Habitat

Brackish meromictic lake sediment

TAS [1]

MIGS-6.3

Salinity

10–40 g NaCl /l

TAS [1]

MIGS-22

Oxygen requirement

obligate anaerobic

TAS [1]

MIGS-15

Biotic relationship

free-living

TAS [1]

MIGS-14

Pathogenicity

non-pathogen

NAS

MIGS-4

Geographic location

Kushiro, Hokkaido, Japan

TAS [1, 5]

MIGS-5

Sample collection

May 2012

TAS [5]

MIGS-4.1

Latitude

42° 58' 20.6" N

TAS [5]

MIGS-4.2

Longitude

144° 24' 6.6" E

TAS [5]

MIGS-4.4

Altitude

NA

 

aEvidence codes - TAS Traceable Author Statement (i.e., a direct report exists in the literature), NAS Non-traceable Author Statement (i.e., not directly observed for the living, isolated sample, but based on a generally accepted property for the species, or anecdotal evidence). NA; not avairable.

Phylogenetic relationship of D. formicivorans strain Pf12BT and all members of the family Desulfomicrobiaceae are shown in the 16S rRNA gene phylogenetic tree (Fig. 2). D. formicivorans strain Pf12BT is assigned to the family Desulfomicrobiaceae but forms a well-separated branch among other cultivated relatives of the same family.
Fig. 2

Phylogenetic tree showing the relationship of D. formicivorans strain Pf12BT to other species of the family Desulfomicrobiaceae. Members of the genus Desulfonatronovibrio were used as the outgroup in this analysis. The tree was constructed by the Maximum-Likelihood method with MEGA version 5.1 [16] based on ClustalX version 2.1 [17] aligned sequences of 16S rRNA gene. Bootstrap values (percentages of 1000 replications) of ≥ 50% are shown at nodes. The presence of sequenced genome is indicated with superscripted “*”

Genome sequencing information

Genome project history

D. formicivorans strain Pf12BT was selected for genome sequencing on the basis of its 16S rRNA gene-based phylogenetic position in the family Desulfomicrobiaceae (Fig. 2). A summary of the genome sequencing project information and its association with MIGS version 2.0 compliance [6] are shown in Table 2. The genome consists of 26 contigs, which has been deposited at DDBJ/EMBL/GenBank under accession number BDFE00000000.
Table 2

Project information

MIGS ID

Property

Term

MIGS 31

Finishing quality

High-quality draft

MIGS-28

Libraries used

TruSeq Nano DNA library prep kit

MIGS 29

Sequencing platforms

Illumina Hiseq paired-end

MIGS 31.2

Fold coverage

370×

MIGS 30

Assemblers

Velvet version 1.2.08

MIGS 32

Gene calling method

Microbial Genome Annotation Pipeline (MiGAP)

 

Locus Tag

BDFE01000001-BDFE01000026

 

Genbank ID

BDFE00000000

 

GenBank Date of Release

June 30, 2016

 

BIOPROJECT

PRJDB4875

MIGS 13

Source Material Identifier

DSM 28890

 

Project relevance

Ecology and evolution

Growth conditions and genomic DNA preparation

D. formicivorans strain Pf12BT (DSM 28890) was grown on bicarbonate-buffered sulfide-reduced medium [7] containing 28 mM sulfate, 10 mM formate and 0.5 g l-1 yeast extract at 45 °C. Genomic DNA was extracted from collected cells using Wizard® genomic DNA purification kit (Promega).

Genome sequencing and assembly

The genome of strain Pf12BT was sequenced using paired-end Illumina sequencing at Hokkaido System Science Co., Ltd. (Japan). From a library with 350 bp inserts, the 10,511,386 reads were generated. After trimming of the reads, a total of 9,393,309 high-quality filtered paired end reads with a hash length of 95 bp were obtained. Reads were assembled de novo using Velvet version 1.2.08 into 26 high quality scaffolds. Gap closing analysis in these scaffolds was performed using Platanus version 1.2.1.

Genome annotation

Draft genome sequences were automatically annotated using the MiGAP [8]. In the pipeline, RNAmmer [9] and tRNAscan-SE [10] were used to identify rRNA and tRNA genes, respectively. MetaGene Annotator [11] was used to predict ORFs likely to encode proteins (CDSs), and functional annotation was performed based on reference databases, including RefSeq, TrEMBL, and COGs. Manual annotation was performed using IMC-GE software (In Silico Biology; Yokohama, Japan). Putative CDSs were confirmed again by a sequence similarity search using the BLASTP tool. Putative CDSs possessing BLASTP matches with more than 70% coverage and 35% identity and E-values less than 1 × e−5 were considered potentially functional genes. When these cut-off values were not satisfied, the CDSs were annotated as hypothetical proteins. Transcription start sites of predicted proteins were corrected based on multiple sequence alignments. If the distance between CDSs was larger than 500 bp, further ORF extraction for coding genes was performed.

The protein-coding genes in the genome were also subjected to analysis on WebMGA [12] for the COGs and Protein family (Pfam) annotations. Transmembrane helices and signal peptide prediction were analyzed using Phobius [13]. CRISPR loci were distinguished using the CRISPR Recognition Tool [14].

Genome properties

The total genome of strain D. formicivorans strain Pf12BT was 3,000,979 bp in size with a GC content of 49.81% (Table 3). It was predicted to contain 2,715 genes including 2,657 protein-coding genes and 58 RNA genes (for tRNA and rRNA). Approximately 83% of the predicted genes were assigned to COG functional categories. The distribution of genes into COGs functional categories is presented in Table 4.
Table 3

Genome statistics

Attribute

Value

% of Total

Genome size (bp)

3,000,979

100.00

DNA coding (bp)

2,596,072

86.51

DNA G + C (bp)

1,494,788

49.81

DNA scaffolds

26

-

Total genes

2,715

100.00

Protein coding genes

2,657

97.86

RNA genes

58

2.14

Pseudo genes

NA

NA

Genes in internal clusters

NA

NA

Genes with function prediction

1888

69.54

Genes assigned to COGs

2255

84.87

Genes with Pfam domains

2110

79.41

Genes with signal peptides

356

13.40

Genes with transmembrane helices

570

21.45

CRISPR repeats

2

0.07

NA, not avairable

Table 4

Number of genes associated with general COG functional categories

Code

Value

%age

Description

J

156

5.75

Translation, ribosomal structure and biogenesis

A

0

0.00

RNA processing and modification

K

102

3.76

Transcription

L

110

4.05

Replication, recombination and repair

B

1

0.04

Chromatin structure and dynamics

D

29

1.07

Cell cycle control, Cell division, chromosome partitioning

V

25

0.92

Defense mechanisms

T

210

7.74

Signal transduction mechanisms

M

169

6.23

Cell wall/membrane biogenesis

N

105

3.87

Cell motility

U

93

3.43

Intracellular trafficking and secretion

O

110

4.05

Posttranslational modification, protein turnover, chaperones

C

222

8.18

Energy production and conversion

G

116

4.27

Carbohydrate transport and metabolism

E

234

8.62

Amino acid transport and metabolism

F

65

2.39

Nucleotide transport and metabolism

H

101

3.72

Coenzyme transport and metabolism

I

51

1.88

Lipid transport and metabolism

P

122

4.50

Inorganic ion transport and metabolism

Q

37

1.36

Secondary metabolites biosynthesis, transport and catabolism

R

258

9.51

General function prediction only

S

168

6.19

Function unknown

-

459

16.91

Not in COGs

Insights from the genome sequence

The draft genome provides interesting phylogenetic and metabolic information, including phylogeny of dsr genes, which are essential for dissimilatory sulfate reduction. The dsrAB genes are frequently used as marker genes to evaluate phylogenetic relationship of sulfate-reducing bacteria, as well as to reveal their diversity and distribution in environments. Phylogenetic analysis based on DsrAB amino acid sequence was performed to disclose the phylogenetic position of D. formicivorans strain Pf12BT among sulfate reducers belonging to the families Desulfovibrionales and Desulfobacterales (Fig. 3). In the resulting phylogenetic tree, D. formicivorans strain Pf12BT was clearly separated from all members of the family Desulfomicrobiaceae . This result partially conflicts with the 16S rRNA gene phylogeny, and this contradiction may represent a new case of lateral gene transfer event which frequently has been found among dissimilatory sulfate-reducing and sulfur-oxidizing bacteria [15].
Fig. 3

Phylogenetic tree based on DsrAB amino acid sequence of D. formicivorans strain Pf12BT and members of the orders Desulfovibrionales and Desulfobacterales. The tree was constructed by the Maximum-Likelihood method with MEGA version 5.1 [16] based on ClustalX version 2.1 [17] aligned protein sequences. Bootstrap values (percentages of 1000 replications) of ≥ 50% are shown at nodes

Conclusions

Draft genome sequence of D. formicivorans strain Pf12BT described here is the first published genome sequence of a member of the genus Desulfoplanes , which is a newly proposed taxon in the family Desulfomicrobiaceae . The genome of the strain Pf12BT consists of 2,657 protein-coding genes and 58 RNA genes. DsrAB phylogenetic tree shows the strain Pf12BT is located in the independent position, which is distant from a cluster of Desulfomicrobium species.

Abbreviations

CRISPR: 

Clustered regularly interspaced short palindromic repeat

Dsr: 

Dissimilatory sulfite reductase

MiGAP: 

Microbial Genome Annotation Pipeline

Declarations

Acknowledgements

This study was supported by a grant-in-aid for Research Fellow of Japan Society for the Promotion Science to MW and JSPS KAKENHI Grant Number 22370005 to MF.

Authors’ contributions

MF and HK designed and supervised the study. MW characterized the strain and carried out all the bioinformatics analysis. MW and HK drafted the manuscript. All authors discussed the data and read and approved the final manuscript.

Competing interests

The authors declare that they have no competing interests.

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Authors’ Affiliations

(1)
The Institute of Low Temperature Science, Hokkaido University
(2)
Postdoctoral Research Fellow of the Japan Society for the Promotion of Science

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Copyright

© The Author(s). 2017