Skip to content

Advertisement

BMC Genomics

What do you think about BMC? Take part in

Open Access

Phylogenetic and comparative genomics of the family Leptotrichiaceae and introduction of a novel fingerprinting MLVA for Streptobacillus moniliformis

  • Tobias Eisenberg1Email author,
  • Ahmad Fawzy1, 2, 5,
  • Werner Nicklas3,
  • Torsten Semmler4 and
  • Christa Ewers5
Contributed equally
BMC Genomics201617:864

https://doi.org/10.1186/s12864-016-3206-0

Received: 9 June 2016

Accepted: 25 October 2016

Published: 3 November 2016

Abstract

Background

The Leptotrichiaceae are a family of fairly unnoticed bacteria containing both microbiota on mucous membranes as well as significant pathogens such as Streptobacillus moniliformis, the causative organism of streptobacillary rat bite fever. Comprehensive genomic studies in members of this family have so far not been carried out. We aimed to analyze 47 genomes from 20 different member species to illuminate phylogenetic aspects, as well as genomic and discriminatory properties.

Results

Our data provide a novel and reliable basis of support for previously established phylogeny from this group and give a deeper insight into characteristics of genome structure and gene functions. Full genome analyses revealed that most S. moniliformis strains under study form a heterogeneous population without any significant clustering. Analysis of infra-species variability for this highly pathogenic rat bite fever organism led to the detection of three specific variable number tandem analysis loci with high discriminatory power.

Conclusions

This highly useful and economical tool can be directly employed in clinical samples without laborious prior cultivation. Our and prospective case-specific data can now easily be compared by using a newly established MLVA database in order to gain a better insight into the epidemiology of this presumably under-reported zoonosis.

Keywords

Next generation sequencingMulti locus variable number tandem repeat analysis (MVLA)PhylogenyTypingFingerprinting Streptobacillus Leptotrichiaceae

Background

The Leptotrichiaceae are a family of underexplored and rarely isolated microorganisms within the phylum Fusobacteria containing both species known from certain pathologies as well as colonising members of the resident microbiota. Many if not all species of the Leptotrichiaceae inhabit the oral cavities, gastrointestinal or urogenital tracts of humans and animals [13]. One of the reasons they are rarely encountered is the obligate anaerobic or capnophilic growth dependence of these fastidious bacteria and the usual presence of a high number of concomitant microorganisms. Some members of this family are well known pathogens, such as Streptobacillus (S.) moniliformis, one of the two causative organisms of the bacterial zoonosis rat bite fever [4]. Recently, a number of novel species have been described, most of which could be attributed to clinical disease [58]. It can also be concluded from numerous phylotypes, Leptotrichiaceae normally colonize mucous membranes [915], but when introduced into new tissue or host sites they are also able to shift their pathogenic potential and cause severe and even life-threatening disease. With increasing availability of next generation sequencing a number of single genomes have been published [6, 1620]. However, almost no comprehensive genomic studies including these microorganisms have been completed, nor have virulence properties been identified in these species. Phylogenetic studies and identifications within the phylum Fusobacteria have been carried out and based on single or multiple gene sequences such as 16S rRNA, 16S–23S rRNA internal transcribed spacer, gyrB, groEL, recA, rpoB, conserved indels and genes for group-specific proteins, 43-kDa outer membrane protein and zinc protease [18, 2130]. In an attempt to characterize different members of this phylum Gupta & Seti proposed various conserved signature indels (CSIs) in amino acid sequences for the Leptotrichiaceae from which three CSIs were found to be specific for this family [31]. On the other hand, no detailed phylogenetic and comparative genome studies dedicated to Leptotrichiaceae have been published up to now. Furthermore, and due to a general paucity of strains and attempts to differentiate members from the same species there is currently no tool available to type isolates in order to prove transmission chains. Our data, presented here, were derived from 46 complete genomes from 20 different taxa of the family Leptotrichiaceae aiming to provide the first such comparative analysis. Our study results confirm the picture of earlier phylogenies from this group that are now based on a larger scale of orthologous genes. We give a surveying insight into the investigated genomes, thereby also including recently described species from this family. With a novel approach it was, furthermore, possible to accurately and unequivocally type isolates of S. moniliformis based on three variable number tandem repeat (VNTR) sequences. With this, we are presenting a culture-independent, species-specific fingerprinting tool in order to type the most important causative organism of rat bite fever for the first time.

Results

Accession numbers

The GenBank/EMBL/DDBJ accession numbers for the genome sequences used in this study are summarized in Table 1.
Table 1

Strains as well as origins, clinical symptoms and host species of the Leptotrichiaceae members used in this study

Strain no.

Strain designation

Species

Year of isolation

Host

Clinic/sample

Country

Strain reference

Genome reference

Accession number

1

DSM 12112T (=ATCC 14647T)

Streptobacillus moniliformis

1925

Human

Rat bite fever

France

[4]

[16]

CP001779.1CP001780.1

2

CIP 55-48

Streptobacillus moniliformis

1947

Mouse

Lymph adenitis

UK

n. d. a.

this study

LWQV00000000

3

ATCC 27747

Streptobacillus moniliformis

1964

Turkey

Septic arthritis

USA

[51]

this study

LWQW00000000

4

NCTC 10773

Streptobacillus moniliformis

1971

Human

Blood culture

UK

n. d. a.

this study

LYRU00000000

5

NCTC 11194

Streptobacillus moniliformis

1977

Human

Rat bite fever

UK

n. d. a.

this study

LWQX00000000

6

IPDH 144/80

Streptobacillus moniliformis

1980

Turkey

Septic arthritis

Germany

n. d. a.

this study

LWQY00000000

7

CIP 81-99

Streptobacillus moniliformis

1981

Human

Blood culture (wild rat bite)

France

n. d. a.

this study

LWSZ00000000

8

AHL 370-4

Streptobacillus moniliformis

1982

Mouse

Ear infection

Australia

n. d. a.

this study

LWTA00000000

9

NCTC 11941

Streptobacillus moniliformis

1983

Human

Haverhill fever

UK

n. d. a.

this study

LXKD00000000

10

IPDH 109/83

Streptobacillus moniliformis

1983

Turkey

Septic arthritis

Germany

n. d. a.

this study

LWTB00000000

11

ATCC 49567

Streptobacillus moniliformis

1989

Mouse

Lymph adenitis

Germany

[52]

this study

LWTC00000000

12

Kun 3 (RIVM)

Streptobacillus moniliformis

1991

Rat

Healthy

The Netherlands

[53]

this study

LWTD00000000

13

ATCC 49940

Streptobacillus moniliformis

1992

Rat

Otitis media

Germany

[54]

this study

LWTE00000000

14

B10/15

Streptobacillus moniliformis

Unknown

Wild rat

Unknown

The Netherlands

n. d. a.

this study

LWTF00000000

15

A378/1

Streptobacillus moniliformis

1995

Wild rat

Vaginal swab

Germany

DKFZ strain collection

this study

LWTG00000000

16

VA11257/2007

Streptobacillus moniliformis

2007

Human (farmer)

Rat bite fever, endocarditis

Germany

[55]

this study

LWTI00000000

17

VK105/14

Streptobacillus moniliformis

2008

Domestic rat

Abscess

Germany

TiHo strain collection

this study

LWTJ00000000

18

B5/1

Streptobacillus moniliformis

2009

Laboratory mouse

After rat bite

Germany

DKFZ strain collection

this study

LXKJ00000000

19

Marseille

Streptobacillus moniliformis

2009

Rat

Rat bite fever

La Réunion

[56]

this study

LXKI00000000

20

IKC1

Streptobacillus moniliformis

n. d. a.

Rat

Oral swab

Japan

[39]

this study

LXKH00000000

21

IKC5

Streptobacillus moniliformis

n. d. a.

Rat

Oral swab

Japan

[39]

this study

LXKG00000000

22

IKB1

Streptobacillus moniliformis

n. d. a.

Rat

Oral swab

Japan

[39]

this study

LXKF00000000

23

TSD4

Streptobacillus moniliformis

n. d. a.

Rat

Oral swab

Japan

[39]

this study

LXKE00000000

24

131000547T (DSM 29248T)

Streptobacillus felis

2013

Cat

Pneumonia

Germany

[5, 7]

[18]

LOHX00000000

25

DSM 26322T (HKU33T)

Streptobacillus hongkongensis

2014

Human

Abscess

Hong Kong

[8]

[18]

LOHY0000000

26

AHL 370-1T

Streptobacillus notomytis

1979

Spinifex hopping mouse

Sepicaemia, cultured from liver tissue

Australia

[57]

[6]

SAMN04038436

27

KWG2

Streptobacillus notomytis

n. d. a.

Rat (Rattus rattus)

Oral swab

Japan

[39]

this study

SAMN04099645

28

KWG24

Streptobacillus notomytis

n. d. a.

Rat (Rattus rattus)

Oral swab

Japan

[39]

this study

SAMN04099670

29

OGS16T

Streptobacillus ratti

n. d. a.

Rat (Rattus rattus)

Oral swab

Japan

[39]

[18]

SAMN04099675

30

CCUG 41628T

Sneathia sanguinegens

1999

Human

Blood

Sweden

[58, 59]

[38]

LOQF00000000

31

Sn35

“Sneathia amnii”

n. d. a.

Human

Vaginal microbiota

n. d. a.

[19]

[19]

NZ_CP011280

32

NCTC 11300T (ATCC 33386T)

Sebaldella termitidis

1962

Termite

Intestine

n. d. a.

[60]

[17]

CP001739

33

DSM 1135 (C-1013-b)

Leptotrichia buccalis

2009

Human

Supragingival calculus

USA

n. d. a.

n. d. a.

CP001685

34

DSM 19756 (LB 57)

Leptotrichia goodfellowii

2013

Human

Prosthetic aortic valve

Germany

n. d. a.

n. d. a.

NZ_AZXW00000000

35

F0264

Leptotrichia goodfellowii

n. d. a.

Human

Oral cavity

n. d. a.

n. d. a.

n. d. a.

NZ_ADAD00000000

36

F0254

Leptotrichia hofstadii

n. d. a.

n. d. a.

n. d. a.

n. d. a.

n. d. a.

n. d. a.

NZ_ACVB00000000

37

DSM 19757

Leptotrichia shahii

2013

Human

Gingivitis

Norway

n. d. a.

n. d. a.

NZ_ARDD00000000

38

DSM 19758

Leptotrichia wadei

2004

Human

Saliva

Norway

[2]

n. d. a.

NZ_ARDS00000000

39

F0279

Leptotrichia wadei

n. d. a.

Human

Subgingival plaque

n. d. a.

n. d. a.

n. d. a.

NZ_AWVM00000000

40

Str. W10393

Leptotrichia sp. oral taxon 212

2015

Human

Oral microbiome project

n. d. a.

n. d. a.

n. d. a.

CP012410

41

Str. W9775

Leptotrichia sp. oral taxon 215

2015

Human

Oral microbiome project

n. d. a.

n. d. a.

n. d. a.

NZ_AWVR00000000

42

Str. F0581

Leptotrichia sp. oral taxon 225

2015

Human

Oral microbiome project

n. d. a.

n. d. a.

n. d. a.

NZ_AWVS00000000

43

Str. F0557

Leptotrichia sp. oral taxon 879

2015

Human

Oral microbiome project

n. d. a.

n. d. a.

n. d. a.

NZ_AWVL00000000

44

CCUG 39713T

Caviibacter abscessus

1998

Guinea pig

Cervical abscess

Sweden

n. d. a.

[38]

LOQG00000000

45

1510011837

Caviibacter abscessus

2015

Guinea pig

Cervical abscess

Germany

[38]

[38]

LOQH00000000

46

AVG2115T

Oceanivirga salmonicida

1992

Atlantic salmon

Septicaemia

Ireland

[32, 61]

[37]

LOQI00000000

47

ATCC 25586

Fusobacterium nucleatum subsp. nucleatum

n. d. a.

Human

Cervico-facial lesion

n. d. a.

n. d. a.

[62]

AE009951

T type strain, n. d. a. no data available, ATCC American Type Culture Collection, Rockville, USA, NCTC National Collection of Type Cultures, London, UK, CIP Collection Institut Pasteur, Paris, France, IPDH Institute for Poultry Diseases, Hannover, Germany, RIVM Rijksinstituut voor Volksgezondheid en Milieuhygiene, Bilthoven, The Netherlands, AHL Animal Health Laboratory, South Perth, Australia, ZfV Zentralinstitut für Versuchstierzucht, Hannover, Germany, DKFZ Deutsches Krebsforschungszentrum, Heidelberg, Germany, TiHo Tierärztliche Hochschule Hannover, Germany, RBF rat bite fever

Phylogenetic analysis based on orthologous genes

To determine the phylogeny within the genus Streptobacillus we aligned the allelic variations of 281 orthologous genes from 29 strains of S. moniliformis, S. ratti, S. notomytis, S. felis and S. hongkongensis which resulted in 57,841 single nucleotide polymorphisms (SNPs). From these SNPs we inferred a maximum likelihood phylogeny showing the distance between the different species within this genus (Fig. 1). To zoom deeper into the phylogeny of the S. moniliformis group we repeated this analyses with 775 orthologous genes present in 23 S. moniliformis strains which resulted in 5,211 SNPs. These SNPs were also used to construct a maximum likelihood phylogeny (Fig. 2).
Fig. 1

Maximum likelihood phylogenetic tree of the genus Streptobacillus (strains 1–29 according to Table 1). The tree is based on 281 orthologous genes including 57,841 SNPs

Fig. 2

Unrooted maximum likelihood phylogenetic tree of 23 Streptobacillus moniliformis strains from this study. The tree is based on 775 orthologous genes including 5,211 SNPs

As shown in the tree, most S. moniliformis strains used for this study are unrelated and form a heterogeneous population without any significant clustering. Solely strains A378/1 and B5/1 that both originate from the same source but without a common epidemiological background were phylogenetically indistinguishable.

Analysis of genomes and protein functions

The genome size in members of the Leptotrichiaceae varies between 1.22 and 4.42 Mbp with Caviibacter (C.) abscessus and Sebaldella (Se.) termitidis being the smallest und largest genomes, respectively. Generally, and with the exception of Sebaldella termitidis, genomes are smaller than 2.45 Mbp. The genera Caviibacter and Sneathia (Sn.) are comparable with respect to genome size (1.22–1.34 Mbp) as are the genera Streptobacillus and Oceanivirga (O.) (1.38–1.90 Mbp). Members of the genus Leptotrichia (L.) are the second largest group with 2.31–2.47 Mbp. A general overview on the genomes of all strains under study is depicted in Table 2. A similar order can be observed with respect to coding DNA sequences (CDS), i.e., C. abscessus and Sneathia spp. possess 1212–1282 CDS, followed by Streptobacillus spp. and O. salmonicida (1293–1679), Leptotrichia spp. (1930–2365) and Sebaldella termitidis (4083). The average percentage of CDS within the whole genome displays a graded distribution within the family: a highly coding group consisting of the genera Caviibacter, Oceanivirga and Sneathia (89–93 %), an intermediate Streptobacillus spp. group (87 %) and a group containing the genera Leptotrichia and Sebaldella (84 %) with lower coding density. Nevertheless, intra-genus variability can be considerably high, the former results can inevitably also be shown for the average gene densities and the average intergenic regions (in parentheses average genes/Mbp; number of intergenic nt): O. salmonicida (1056; 79), C. abscessus (996; 76), Sneathia spp. (989; 84), Streptobacillus spp. (987; 115), Leptotrichia spp. (967; 144) and Sebaldella (936; 149). An organization of the genomes under study into clusters of orthologous groups (COGs) is depicted in Additional files 1 and 2 and shows, however, high intra-species as well as inter-species variations. On a generic level, gene contents of COG classes J, L, D and F are inversely correlated with increasing genome size, whereas COG classes K, N, T and Q are positively correlated (see Additional files 1 and 2).
Table 2

Analysis of genome data as well as predictions of coding regions of the Leptotrichiaceae members used in this study

Strain no.

Organism

Approx. genome size (nt)

CDSa

rRNA

tRNAb

% GCc

Total DNA coding regions (nt)

Total non-coding regions (nt)

Coding genome space (%)

Average gene density (genes/Mbp)

Average inter-genic region (nt)

1

S. moniliformis

1673280

1568

16

39

26.3

1556870

116410

93

937

74

2

S. moniliformis

1678906

1658

12

37

26.1

1508835

170071

89

988

103

3

S. moniliformis

1684459

1591

14

35

26.1

1486041

198418

87

945

125

4

S. moniliformis

1897024

2244

9

43

28.9

1651665

245359

85

1183

109

5

S. moniliformis

1712153

1764

3

38

26.1

1542831

169322

89

1030

96

6

S. moniliformis

1668382

1615

13

36

26.1

1484745

183637

88

968

114

7

S. moniliformis

1686977

1543

12

35

26.4

1449924

237053

84

915

154

8

S. moniliformis

1598404

1608

14

38

25.9

1470174

128230

91

1006

80

9

S. moniliformis

1689124

1675

4

36

26.1

1399686

289438

79

992

173

10

S. moniliformis

1756513

1765

14

37

26.1

1559103

197410

87

1005

112

11

S. moniliformis

1763717

1621

9

35

26.1

1488168

275549

81

919

170

12

S. moniliformis

1518628

1540

12

33

25.9

1442043

76585

95

1014

50

13

S. moniliformis

1689360

1765

5

36

26.1

1526748

162612

89

1045

92

14

S. moniliformis

1674237

1597

13

37

26.2

1477515

196722

87

954

123

15

S. moniliformis

1667701

1692

14

36

26.0

1518810

148891

90

1015

88

16

S. moniliformis

1690579

1538

16

37

26.1

1468143

222436

85

910

145

17

S. moniliformis

1608659

1507

22

34

26.2

1433763

174896

88

937

116

18

S. moniliformis

1497161

1644

8

36

25.8

1322022

175139

87

1098

107

19

S. moniliformis

1696954

1774

5

38

26.1

1521612

175342

88

1045

99

20

S. moniliformis

1696554

1688

17

37

26.0

1509528

187026

88

995

111

21

S. moniliformis

1792325

1664

16

42

26.2

1550631

241694

84

928

145

22

S. moniliformis

1759287

1737

13

43

25.9

1566621

192666

88

987

111

23

S. moniliformis

1608076

1559

10

35

26.0

1445580

162496

89

969

104

24

S. felis

1610666

1754

3

37

26.4

1450014

160652

89

1089

92

25

S. hongkongensis

1543001

1485

14

35

26.1

1324059

218942

83

962

147

26

S. notomytis

1762984

1773

9

43

28.1

1511157

251827

83

1006

142

27

S. notomytis

1426245

1349

8

40

26.4

1257996

168249

87

946

125

28

S. notomytis

1384502

1341

19

39

26.3

1256817

127685

90

969

95

29

S. ratti

1499353

1411

11

39

25.9

1318767

180586

86

941

128

30

Sneathia sanguinegens

1300753

1329

2

34

26.7

1214541

86212

93

1022

65

31

“Sn. amnii”

1339284

1282

 

34

28.3

1207722

131562

89

957

103

32

Sebaldella termitidis

4418842

4135

13

40

33.5

3802074

616768

84

936

149

33

Leptotrichia buccalis

2465610

2299

15

46

29.6

2062809

402801

80

932

175

34

L. goodfellowii

2281162

2241

7

39

31.6

2045213

235949

88

982

105

35

L. goodfellowii

2287284

2373

3

39

31.5

2055020

232264

89

1037

98

36

L. hofstadii

2453253

2720

13

47

30.8

2059248

394005

81

1109

145

37

L. shahii

2144606

1969

10

41

29.5

1812950

331656

82

918

168

38

L. wadei

2316529

2139

11

42

29.3

1973929

342600

83

923

160

39

L. wadei

2353455

2212

3

27

29.2

2008568

344887

83

940

156

40

Leptotrichia sp. oral taxon 212

2444904

2231

14

43

31.4

2146482

298422

86

936

130

41

Leptotrichia sp. oral taxon 215

2308492

2195

3

34

31.4

2039067

269425

87

951

123

42

Leptotrichia sp. oral taxon 225

2400083

2306

3

24

29.6

2061283

338800

84

961

147

43

Leptotrichia sp. oral taxon 879

2415750

2361

4

25

29.6

2026284

389466

81

977

165

44

C. abscessus

1219935

1198

  

26.5

1131456

88479

92

982

74

45

C. abscessus

1304155

1316

4

35

26.4

1201320

102835

91

1009

78

46

O. salmonicida

1769081

1869

2

38

25.4

1621182

147899

91

1056

79

47

Fusobacterium nucleatum (outgroup)

2174500

2022

15

47

27.2

1937724

236776

88

930

117

aCDS: DNA coding sequences; btRNA: transfer ribonucleic acid; cGC: guanine-cytosine content

Multiple-Locus Variable number tandem repeat Analysis (MLVA)

In silico VNTR analysis

Under default conditions, 127 repeats were identified by the tandem repeat finder. For further analysis, the three most variable VNTRs were identified according to the degree of variability of allele types identified by alignment analysis (Table 3). These three allelic loci were only present in S. moniliformis and thus proved to be specific for this microorganism (all other members of the Leptotrichiaceae were negative). The combination of the three loci yielded a high discriminatory index (0.94296 DI; Table 4).
Table 3

Streptobacillus moniliformis specific Variable Number of Tandem Repeat (VNTR) primer sequences used in this study

Primer ID

VNTR positiona

Repeat size in nt (identity in %)

Sequence (5–3)

PCR product size (bp)

VNTR_Sm1

1576120 - 1576145

3 (100)

TCA TTT ACT CAC CCT AGT AGT GGT

210

CCA GTT GAA TAT AAG CTT GCT ATG G

VNTR_Sm2

1182890 - 1182907

6 (100)

TGG AAC TGT TTG TTG AGT ATT TCC A

298

AGG GAC AGA TGT TCA ATT TGT GTA

VNTR_Sm3

284997 - 285268

36 (91)

TAC GCT GTA GGG TTG AAC GG

830

ACA GTT TGA GCA CGT CTT AAT CC

Primers were designed with Geneious (v. 8.1.3; Biomatters, Auckland, NZ) [43] and to be complementary to VNTR flanking regions that were conserved among genomes; aaccording to the S. moniliformis DSM 12112T genome (CP001779.1)

Table 4

VNTR allele types of the Streptobacillus moniliformis strains used in this study

Isolate ID

VNTR_Sm1a

VNTR_Sm2

VNTR_Sm3b

Allele code

DSM 12112 T

9

3

16

LHL1

CIP 55-48

7

3

16

LHL10

ATCC 27747

10

3

16

LHL4

NCTC 10773

8

4

17

LHL15

NCTC 11194

6

3

17

LHL16

IPDH 144/80

6

3

16

LHL5

CIP 81-99

7

3

16

LHL10

AHL 370-4

7

2

15

LHL3

NCTC 11941

6

3

18

LHL11

IPDH 109/83

6

3

16

LHL5

ATCC 49567

6

3

16

LHL5

Kun 3 (RIVM)

6

3

18

LHL11

ATCC 49940

6

3

14

LHL6

B10/15

6

4

15

LHL7

A378/1

8

5

16

LHL2

VA11257/2007

6

3

16

LHL5

VK105/14

8

3

16

LHL13

B5/1

8

5

16

LHL2

Marseille

6

4

14

LHL14

IKC1

6

3

15

LHL8

IKC5

5

3

15

LHL9

IKB1

6

3

16

LHL5

TSD4

11

3

18

LHL12

A40-13 c

11

2

17

LHL17

Bold rows represent strains used for a PCR-based validation of in silico identified VNTR allele types (underlined alleles were not found in silico and only identified after PCR amplification); a in order to fit requirements of the database, the repeat copy numbers at locus VNTR_Sm1 have been rounded up to receive integer values (e.g., 9 instead of 8.7); bwhile the repeat copy numbers at locus VNTR_Sm3 have been rounded up to the next half-value and doubled to receive integer values (e.g., 15 instead of 7.2); T: type strain; cstrain was only used for validation (no complete genome available)

PCR-based validation of in silico results

The absence of the calculated VNTR loci could also be proven by polymerase chain reaction (PCR) in all Leptotrichiaceae members other than S. moniliformis (data not shown). Contrarily, each of the ten S. moniliformis strains exhibited a specific band corresponding to their predicted tandem repeats pattern. Analysis of the sequenced PCR products confirmed the allele type allocation determined in silico (Table 4). VNTR_Sm1 alleles of two isolates, which were not found in silico, were successfully assigned (Table 4). Re-calculation revealed a DI of 0.9529 after including these two isolates, as well as one isolate for which no genome data was available. In order to facilitate comparisons of results in future studies, every genotype (from the allele types of the three loci) was expressed as a specific allele combination resulting in a specific allele code (Table 4). An online database dedicated to MLVA results of S. moniliformis has been established on the webserver of University Paris-Saclay, Orsay, France (http://microbesgenotyping.i2bc.paris-saclay.fr/databases/public) which is open to future entries and strain comparisons.

Discussion

Members of the Leptotrichiaceae are rarely encountered microorganisms, a phenomenon that seems to be highly dependent on difficulties with cultivation. With the availability of molecular methods in this field the number of findings and frequencies has significantly increased [1015, 3236]. On the other hand, we still need deeper insight into the genomes of this group. In particular, the mechanisms involved in pathogenesis and virulence of pathogenic species are completely unexplored. We have undertaken a first step into this direction by analysing a broad spatio-temporal collection of strains, thereby including especially species with regular evidence for pathologies. Firstly, the large dataset from this study has been utilized for the confirmation of our phylogenetic picture from earlier studies [18, 30, 37, 38]. An intra-genus phylogeny that was based on 775 orthologous genes revealed a very similar picture to previous studies involving only four selected functional genes (Figs. 1 and 2). Conversely and in contrast to almost identical average nucleotide identity (ANI) values [30], full genome analyses revealed a high level of heterogeneity for all but two strains (no. 15 and 18) of S. moniliformis without any significant clustering. This is, albeit, not surprising, because the present study included a large spatio-temporal collection of 23 S. moniliformis strains that have been isolated over a period of 90 years from at least five different host species and from almost all subcontinents. We were also able to display the three predicted Leptotrichiaceae specific CSIs of MreB/MrI (2 aa deletion), AlaS and RecA (5 and 2 aa insertions, respectively) in all of our genomes as well as in the recently described members of the family (data not shown) [31].

Genome size dependent gene content has been described and could also be confirmed for the genomes from this study [19]. With increasing genome size gene contents of COG classes J, L, D and F involved in DNA replication, cell cycle regulation and protein translation are inversely correlated, whereas COG classes K, N, T and Q involved in transcription, signal transduction, cell motility and the biochemistry of secondary metabolites are positively correlated (see Additional files 1 and 2). This makes sense when essential gene functions are preserved in smaller genomes and less important gene functions which are dispensable or can be ‘outsourced’ to the host, are lost [19]. On first impression the group of S. moniliformis strains is highly similar as can be concluded from related morphological and phenotypical properties and also from their high intra-species ANI of 98.5–99.3 % (cf. Table S2 in [30]). Based on data from this study very similar COG classes were also observed within this group (see Additional files 1 and 2), but differences in coding densities suggested, on the other hand, remarkable discrepancies. Fuelled by the idea that these discrepancies could, furthermore, be utilized with respect to epidemiology, we have developed a specific MLVA typing scheme for the major pathogen from this group, S. moniliformis, and the causative organism of rat bite fever. This scheme proved to be sufficient in unequivocally typing all 23 S. moniliformis strains under study plus one additional isolate with high discriminatory power (0.9529 DI). Interestingly, only four allele codes (genotypes; LHL2, LHL5, LHL10 and LHL11) were found more than once among isolates (Table 4). At least for LHL2 isolates, a connection could be pursued in that both isolates have been stored in the same strain collection, although a direct transmission could not be proven. To check the clonality of isolates belonging to these four genotypes we have investigated further loci with high discriminatory potential, i.e., the clustered regularly interspaced short palindromic repeats (CRISPR) region known to occur in S. moniliformis (http://crispr.u-psud.fr/cgi-bin/crispr/SpecieProperties.cgi?Taxon_id=519441). In contrast to all other allele codes (LHL5, LHL10, LHL11), both strains (no. 15 and 18) belonging to the allele code LHL2 indeed shared an identical CRISPR region, thereby pointing towards a clonal relation of these two isolates (data not shown) as could also be concluded from the phylogenetic tree (Fig. 2). Due to its length of up to approximately 3,000 nucleotides and its high level of heterogeneity the CRISPR region seems, on the other hand, presently not very well suited as a direct typing tool, but could be useful in certain situations to confirm or negate clonality of strains. A second advantage of the MLVA method described in this study is that it can effectively be pursued directly from the original matrix (e.g., a mouth microbiota swab and a clinical sample) without prior cultivation of the organism, which offers the possibility to better understand transmission chains in the future. This seems to be especially relevant since established PCR assays are not species specific, but limited to genus level specificity [37, 39, 40]. The majority of diagnoses of rat bite fever cases in the recently published literature relies only on partial 16S rRNA gene sequence analysis that may – in the light of very similar novel Streptobacillus spp. that also colonize rats – be quite uncertain for proper pathogen identification [41]. Hopefully, the newly established MLVA database will help to clarify regional infectious clusters and confirm transmission of certain lineages.

Conclusion

We have undertaken a first analysis of Leptotrichiaceae genomes using a large spatiotemporal collection of strains also including novel members of this group. Our dataset unveiled a first insight into characteristics founding a stable phylogeny, genome structure and COG classes. Beside apparent intra-species similarities we have detected also genetic heterogeneities that provided a basis for fingerprinting the most relevant pathogen from this group, the rat bite fever organism, S. moniliformis. This highly useful and economical tool can be directly used from clinical samples without ambitious prior cultivation and with high discriminatory power. Our data form the basis for a newly established MLVA database that provides the opportunity to store and compare isolate-specific information in future cases with this neglected zoonosis.

Methods

Generation of genomic data

Twenty-two strains of S. moniliformis were sequenced in this study, ten strains were taken from previous publications of our group and 15 strains were descended from other projects (Table 1). Genomic DNA was extracted from a 72 h bacterial culture with a commercial kit according to the manufacturer’s instructions (MasterPure™ Complete DNA and RNA Purification Kit, Epicentre, distributed by Biozym Scientific, Hessisch Oldendorf, Germany). Whole genome sequencing of the strains was performed on an Illumina MiSeq with v3 chemistry resulting in 300 bp paired end reads and a coverage of greater than 90×. Quality trimming and de novo assembly was performed with CLC Genomics Workbench, Version 7.5 (CLC Bio, Aarhus, Denmark). For automatic annotation we used the RAST Server: Rapid Annotations using Subsystems Technology [42]. Data from further relevant reference genomes from the Leptotrichiaceae were also utilized and obtained from the National Center for Biotechnology Information (NCBI) database (http://www.ncbi.nlm.nih.gov). Sequence analyses and genome calculations as well as oligonucleotide primer generation were carried out with Geneious (v. 8.1.3; Biomatters, Auckland, NZ) [43]. Table 1 depicts the set of strains and reference genomes used for this study.

Phylogenetic analysis based on orthologous genes

The determination of the maximum common genome (MCG) alignment was done comprising those genes present in all genomes considered for comparison [44]. Based on the parameters sequence similarity (minimum 70 %) and coverage (minimum 90 %) the genes were clustered and those genes that were present in each genome, fulfilling the threshold parameters were defined as MCG. This resulted in 281 orthologous genes for the comparison of 29 strains of S. moniliformis, S. ratti, S. notomytis, S. felis and S. hongkongensis and in 775 orthologous genes for the comparison within 23 strains of S. moniliformis only.

The following extraction of the allelic variants of these genes from all genomes was performed by a blast based approach after which they were aligned individually for each gene and concatenated which resulted in an alignment of 219,961 bp for the 29 strains and of 546,508 bp for the 23 S. moniliformis strains [45].

This alignment was used to generate a phylogenetic tree with randomized axelerated maximum likelihood (RAxML) 8.1 [46] using a General Time Reversible model and gamma correction for among site rate variation.

Analysis of genomes and protein functions

Genes were predicted with Prodigal [47] and assigned to COGs with the NCBI’s Conserved Domain Database [48].

Multiple-Locus Variable number tandem repeat Analysis (MLVA)

In silico VNTR analysis

The complete genome sequence of the S. moniliformis type strain DSM12112T (accession number CP001779.1) was used to search for potential VNTRs using a tandem repeat finder web tool (http://tandem.bu.edu/trf/trf.basic.submit.html). We focused our search on repeats that were characterized by high purity, large size, and/or large number of repeat copies [49]. Repeats of interest were aligned against a set of available genomes depicted in Table 1 using Geneious and allele types were determined as shown in repeat copy numbers. The DI was calculated for a combination of three most variable VNTRs using an online discriminatory power calculator (http://insilico.ehu.es/mini_tools/discriminatory_power/).

PCR-based validation of in silico results

Ten S. moniliformis strains (strain nos. 1, 2, 3, 12, 14, 15, 21, 22 and 23 according to Table 1 plus strain A40-13 for which complete genomic data were not available) as well as all accessible members of the Leptotrichiaceae other than S. moniliformis were used for validation. DNA was extracted from respective isolates (2–3 colonies) by boiling in 100 μL distilled water for 20 min (min.) followed by centrifugation at 20,817 × g for 5 min. The 20 μL final PCR reaction contained 10 μL of Hotstar Taq MasterMix (Qiagen, Hilden, Germany), 1 μL of each forward and reverse primer (10 pmol/μL) (TIB MOLBIOL, Berlin, Germany) (Table 3), 6 μL DNase free PCR grade water (Qiagen), and 2 μL of the extracted DNA. PCR conditions were as following: 1× (95 °C, 15 min), 40x (94 °C, 30 s; 58 °C, 30 s; 72 °C, 30 s), 1× (72 °C, 10 min). PCR products were stained with ethidium bromide in a 2 % agarose gel (100 V for 1.5 h) and then analyzed with a gel documentation system (BioDoc-It, UVP, UK). The PCR amplicons were purified using MicroElute DNA Cycle-Pure Kit (OMEGA bio-tek, Norcross, USA) and sequenced at Seqlab-Microsynth laboratories (Göttingen, Germany). All sequences were analyzed by tandem repeat finder web tool and/or BLASTN 2.3.1+ [50] hosted by NCBI website and compared to the in silico results.

Abbreviations

AHL: 

Animal Health Laboratory South Perth, Australia

ANI: 

Average nucleotide identity

ATCC: 

American Type Culture Collection Rockville, USA

C.: 

Caviibacter

°C: 

Degrees Celsius

CDS: 

Coding DNA sequences

CIP: 

Collection Institut Pasteur Paris, France

COG: 

Cluster of orthologous groups

CRISPR: 

Clustered regularly interspaced short palindromic repeat

CSI: 

Conserved signature indel

DDBJ: 

DNA Data Bank of Japan

DI: 

Discriminatory index

DNA: 

Desoxyribonucleic acid

DKFZ: 

Deutsches Krebsforschungszentrum Heidelberg, Germany

EMBL: 

European Molecular Biology Laboratory

Fig.: 

Figure

g

Gravity

GC: 

Guanine-cytosine content

h: 

Hour

IPDH: 

Institute for Poultry Diseases Hannover, Germany

kDa: 

kilo Dalton

L.

Leptotrichia

LHL: 

Landesbetrieb Hessisches Landeslabor

Mbp: 

Mega base pairs

MCG: 

Maximum common genome

min: 

minute

MVLA: 

Multi locus variable number tandem repeat analysis

μL: 

micro liter

NCBI: 

National Center for Biotechnology Information

NCTC: 

National Collection of Type Cultures London, UK

n. d. a.: 

no data available

nt: 

nucleotides

O.: 

Oceanivirga

PCR: 

Polymerase chain reaction

RAST: 

Rapid annotations using subsystems technology

RaxML: 

Randomized axelerated maximum likelihood

RBF: 

Rat bite fever

RIVM: 

Rijksinstituut voor Volksgezondheid en Milieuhygiene Bilthoven, The Netherlands

rRNA: 

ribosomal ribonucleic acid

S.

Streptobacillus

Se.

Sebaldella

sec.: 

second

Sn.

Sneathia

SNPs: 

Single nucleotide polymorphisms

T

Type strain

TiHo: 

Tierärztliche Hochschule Hannover, Germany

tRNA: 

transfer ribonucleic acid

VNTR: 

Variable number tandem repeat

ZfV: 

Zentralinstitut für Versuchstierzucht, Hannover, Germany

Declarations

Acknowledgement

We thank Ulrike Kling, Asmahan Omar, Anna Mohr, Katharina Engel, Mersiha Curić, Jens Heinbächer, Ursula Leidner, Andrea Erles-Kemna and Bernhard Berkus for excellent technical assistance and Barbara Gamb for making even the most exotic manuscripts available. We are greatly acknowledging the support of Walter Geißdörfer (Erlangen), Judith Rohde (Hannover), Bernard La Scola (Marseille), Koichi Imaoka (Tokyo) and Nobuhito Hayashimoto (Tokyo) for providing strains or DNA of strains no. 17, 18, 19, 21–28 and A40-13, respectively.

Funding

We declare that none of the authors has received funding for this work.

Availability of data and materials

The GenBank/EMBL/DDBJ accession numbers for the genome sequences used in this study are summarized in Table 1. Phylogenetic datasets generated during and analysed during the current study are available in the Dryad Digital Repository, http://dx.doi.org/10.5061/dryad.1q7q4 [45].

Authors’ contributions

TE, CE and TS conceived the study. TE, AF, WN, and TS carried out diagnostics and experiments. TE, CE and TS conducted the data analysis. TE wrote the manuscript and all the authors read and approved the final manuscript.

Competing interests

The authors declare that they have no competing interests.

Consent for publication

Not applicable.

Ethics approval and consent to participate

“Not applicable” (This manuscript does not contain any human or animal participants, human or animal data, or human or animal tissue and therefore does not require a statement on ethics approval and consent.)

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

Authors’ Affiliations

(1)
Abteilung Veterinärmedizin, Landesbetrieb Hessisches Landeslabor (LHL)
(2)
Department of Medicine and Infectious Diseases, Faculty of Veterinary Medicine, Cairo University
(3)
Deutsches Krebsforschungszentrum
(4)
Robert Koch-Institut
(5)
Institut für Hygiene und Infektionskrankheiten der Tiere, Justus-Liebig-Universität Giessen

References

  1. Gaastra W, Boot R, Ho HT, Lipman LJ. Rat bite fever. Vet Microbiol. 2009;133(3):211–28.View ArticlePubMedGoogle Scholar
  2. Eribe ER, Paster BJ, Caugant DA, Dewhirst FE, Stromberg VK, Lacy GH, Olsen I. Genetic diversity of Leptotrichia and description of Leptotrichia goodfellowii sp. nov., Leptotrichia hofstadii sp. nov., Leptotrichia shahii sp. nov. and Leptotrichia wadei sp. nov. Int J Syst Evol Microbiol. 2004;54(Pt 2):583–92.View ArticlePubMedGoogle Scholar
  3. Harwich Jr MD, Serrano MG, Fettweis JM, Alves JM, Reimers MA, Buck GA, Jefferson KK. Genomic sequence analysis and characterization of Sneathia amnii sp. nov. BMC Genomics. 2012;13 Suppl 8:S4.View ArticlePubMedPubMed CentralGoogle Scholar
  4. Levaditi C, Nicolau S, Poincloux P. Sur le rôle étiologique de Streptobacillus moniliformis (nov. spec.) dans l’érythème polymorphe aigu septicémique. C R Acad Sci. 1925;180:1188–90.Google Scholar
  5. Eisenberg T, Glaeser S, Nicklas W, Mauder N, Contzen M, Aledelbi K, Kämpfer P. Streptobacillus felis sp. nov. isolated from a cat with pneumonia. Int J Syst Evol Microbiol. 2015;65(7):2172–8.View ArticlePubMedGoogle Scholar
  6. Eisenberg T, Glaeser SP, Ewers C, Semmler T, Nicklas W, Rau J, Mauder N, Hofmann N, Imaoka K, Kimura M, et al. Streptobacillus notomytis sp. nov. isolated from a spinifex hopping mouse (Notomys alexis THOMAS, 1922), and emended description of Streptobacillus Levaditi et al. 1925, Eisenberg et al. 2015 emend. Int J Syst Evol Microbiol. 2015;65(12):4823–9.View ArticlePubMedGoogle Scholar
  7. Eisenberg T, Nesseler A, Nicklas W, Spamer V, Seeger H, Zschöck M. Streptobacillus sp. isolated from a cat with pneumonia. J Clin Microbiol Case Reports. 2014;2014:1–7.Google Scholar
  8. Woo PC, Wu AK, Tsang CC, Leung KW, Ngan AH, Curreem SO, Lam KW, Chen JH, Chan JF, Lau SK. Streptobacillus hongkongensis sp. nov., isolated from patients with quinsy and septic arthritis, and emended descriptions of the genus Streptobacillus and the species Streptobacillus moniliformis. Int J Syst Evol Microbiol. 2014;64(Pt 9):3034–9.View ArticlePubMedGoogle Scholar
  9. Bik EM, Rohlik CM, Chow E, Carlin KP, Jensen ED, Venn-Watson S, Relman DA. Indigenous microbiota of marine mammals. In: 13th International Symposium on Microbial Ecology. Seattle: International Society for Microbial Ecology; 2010. http://www.isme-microbes.org/isme13.Google Scholar
  10. Chaves-Moreno D, Plumeier I, Kahl S, Krismer B, Peschel A, Oxley AP, Jauregui R, Pieper DH. The microbial community structure of the cotton rat nose. Environ Microbiol Rep. 2015;7(6):929–35.View ArticlePubMedGoogle Scholar
  11. Dewhirst FE, Klein EA, Thompson EC, Blanton JM, Chen T, Milella L, Buckley CM, Davis IJ, Bennett ML, Marshall-Jones ZV. The canine oral microbiome. PLoS One. 2012;7(4):e36067.View ArticlePubMedPubMed CentralGoogle Scholar
  12. Hullar MA, Lancaster SM, Li F, Tseng E, Beer K, Atkinson C, Wahala K, Copeland WK, Randolph TW, Newton KM, et al. Enterolignan-producing phenotypes are associated with increased gut microbial diversity and altered composition in premenopausal women in the United States. Cancer Epidemiol Biomarkers Prev. 2015;24(3):546–54.View ArticlePubMedGoogle Scholar
  13. Strong T, Dowd S, Gutierrez AF, Coffman J. Amplicon pyrosequencing of wild duck eubacterial microbiome from a fecal sample reveals numerous species linked to human and animal diseases. Research. 2013;2(224):1–7. [v1; ref status: awaiting peer review, http://f1000r.es/1yy]. F1000.Google Scholar
  14. Xenoulis PG, Palculict B, Allenspach K, Steiner JM, Van House AM, Suchodolski JS. Molecular-phylogenetic characterization of microbial communities imbalances in the small intestine of dogs with inflammatory bowel disease. FEMS Microbiol Ecol. 2008;66(3):579–89.View ArticlePubMedGoogle Scholar
  15. Bik EM, Costello EK, Switzer AD, Callahan BJ, Holmes SP, Wells RS, Carlin KP, Jensen ED, Venn-Watson S, Relman DA. Marine mammals harbor unique microbiotas shaped by and yet distinct from the sea. Nat Commun. 2016;7:10516.View ArticlePubMedPubMed CentralGoogle Scholar
  16. Nolan M, Gronow S, Lapidus A, Ivanova N, Copeland A, Lucas S, Del Rio TG, Chen F, Tice H, Pitluck S, et al. Complete genome sequence of Streptobacillus moniliformis type strain (9901). Stand Genomic Sci. 2009;1(3):300–7.PubMedPubMed CentralGoogle Scholar
  17. Harmon-Smith M, Celia L, Chertkov O, Lapidus A, Copeland A, Glavina Del Rio T, Nolan M, Lucas S, Tice H, Cheng JF, et al. Complete genome sequence of Sebaldella termitidis type strain (NCTC 11300). Stand Genomic Sci. 2010;2(2):220–7.View ArticlePubMedPubMed CentralGoogle Scholar
  18. Eisenberg T, Imaoka K, Kimura M, Glaeser SP, Ewers C, Semmler T, Rau J, Nicklas W, Kämpfer P. Streptobacillus ratti sp. nov., isolated from a black rat (Rattus rattus). Int J Syst Evol Microbiol. 2016;66(4):1620–6.View ArticlePubMedGoogle Scholar
  19. Harwich Jr MD, Serrano MG, Fettweis JM, Alves JM, Reimers MA, Vaginal Microbiome Consortium (additional members), Buck GA, Jefferson KK. Genomic sequence analysis and characterization of Sneathia amnii sp. nov. BMC Genomics. 2012;13 Suppl 8:S4.View ArticlePubMedPubMed CentralGoogle Scholar
  20. Ivanova N, Gronow S, Lapidus A, Copeland A, Glavina Del Rio T, Nolan M, Lucas S, Chen F, Tice H, Cheng JF, et al. Complete genome sequence of Leptotrichia buccalis type strain (C-1013-b). Stand Genomic Sci. 2009;1(2):126–32.View ArticlePubMedPubMed CentralGoogle Scholar
  21. Woo PC, Wong SS, Teng JL, Leung KW, Ngan AH, Zhao DQ, Tse H, Lau SK, Yuen KY. Leptotrichia hongkongensis sp. nov., a novel Leptotrichia species with the oral cavity as its natural reservoir. J Zhejiang Univ Sci B. 2010;11(6):391–401.View ArticlePubMedPubMed CentralGoogle Scholar
  22. Conrads G, Claros MC, Citron DM, Tyrrell KL, Merriam V, Goldstein EJ. 16S-23S rDNA internal transcribed spacer sequences for analysis of the phylogenetic relationships among species of the genus Fusobacterium. Int J Syst Evol Microbiol. 2002;52(Pt 2):493–9.View ArticlePubMedGoogle Scholar
  23. Sun D, Zhang H, Lv S, Wang H, Guo D. Identification of a 43-kDa outer membrane protein of Fusobacterium necrophorum that exhibits similarity with pore-forming proteins of other Fusobacterium species. Res Vet Sci. 2013;95(1):27–33.View ArticlePubMedGoogle Scholar
  24. Kim HS, Lee DS, Chang YH, Kim MJ, Koh S, Kim J, Seong JH, Song SK, Shin HS, Son JB, et al. Application of rpoB and zinc protease gene for use in molecular discrimination of Fusobacterium nucleatum subspecies. J Clin Microbiol. 2010;48(2):545–53.View ArticlePubMedGoogle Scholar
  25. Shah HN, Olsen I, Bernard K, Finegold SM, Gharbia S, Gupta RS. Approaches to the study of the systematics of anaerobic, gram-negative, non-sporeforming rods: current status and perspectives. Anaerobe. 2009;15(5):179–94.View ArticlePubMedGoogle Scholar
  26. Strauss J, White A, Ambrose C, McDonald J, Allen-Vercoe E. Phenotypic and genotypic analyses of clinical Fusobacterium nucleatum and Fusobacterium periodonticum isolates from the human gut. Anaerobe. 2008;14(6):301–9.View ArticlePubMedGoogle Scholar
  27. Jin J, Haga T, Shinjo T, Goto Y. Phylogenetic analysis of Fusobacterium necrophorum, Fusobacterium varium and Fusobacterium nucleatum based on gyrB gene sequences. J Vet Med Sci. 2004;66(10):1243–5.View ArticlePubMedGoogle Scholar
  28. Jalava J, Eerola E. Phylogenetic analysis of Fusobacterium alocis and Fusobacterium sulci based on 16S rRNA gene sequences: proposal of Filifactor alocis (Cato, Moore and Moore) comb. nov. and Eubacterium sulci (Cato, Moore and Moore) comb. nov. Int J Syst Bacteriol. 1999;49(Pt 4):1375–9.View ArticlePubMedGoogle Scholar
  29. Lawson PA, Gharbia SE, Shah HN, Clark DR, Collins MD. Intrageneric relationships of members of the genus Fusobacterium as determined by reverse transcriptase sequencing of small-subunit rRNA. Int J Syst Bacteriol. 1991;41(3):347–54.View ArticlePubMedGoogle Scholar
  30. Eisenberg T, Nicklas W, Mauder N, Rau J, Contzen M, Semmler T, Hofmann N, Aledelbi K, Ewers C. Phenotypic and genotypic characteristics of members of the genus Streptobacillus. PLoS One. 2015;10(8):e0134312.View ArticlePubMedPubMed CentralGoogle Scholar
  31. Gupta RS, Sethi M. Phylogeny and molecular signatures for the phylum Fusobacteria and its distinct subclades. Anaerobe. 2014;28:182–98.View ArticlePubMedGoogle Scholar
  32. Palmer R, Drinan E, Murphy T. A previously unknown disease of farmed Atlantic salmon: pathology and establishment of bacterial aetiology. Dis Aquat Org. 1994;19:7–14.View ArticleGoogle Scholar
  33. Wouters EG, Ho HT, Lipman LJ, Gaastra W. Dogs as vectors of Streptobacillus moniliformis infection? Vet Microbiol. 2008;128(3–4):419–22.View ArticlePubMedGoogle Scholar
  34. Swartz JD, Lachman M, Westveer K, O’Neill T, Geary T, Kott RW, Berardinelli JG, Hatfield PG, Thomson JM, Roberts A, et al. Characterization of the vaginal microbiota of ewes and cows reveals a unique microbiota with low levels of lactobacilli and near-neutral pH. Front Vet Sci. 2014;1:19.View ArticlePubMedPubMed CentralGoogle Scholar
  35. Kong HH, Oh J, Deming C, Conlan S, Grice EA, Beatson MA, Nomicos E, Polley EC, Komarow HD, Program NCS, et al. Temporal shifts in the skin microbiome associated with disease flares and treatment in children with atopic dermatitis. Genome Res. 2012;22(5):850–9.View ArticlePubMedPubMed CentralGoogle Scholar
  36. Eisenberg T, Glaeser SP, Kämpfer P, Schauerte N, Geiger C. Root sepsis associated with insect-dwelling Sebaldella termitidis in a lesser dwarf lemur (Cheirogaleus medius). Antonie Van Leeuwenhoek. 2015;108(6):1373–82.View ArticlePubMedGoogle Scholar
  37. Eisenberg T, Kämpfer P, Ewers C, Semmler T, Glaeser SP, Collins E, Ruttledge M, Palmer R. Oceanivirga salmonicida gen. nov. sp. nov., a novel member from the Leptotrichiaceae isolated from Atlantic salmon (Salmo salar). Int J Syst Evol Microbiol. 2016;66:2429–37.View ArticlePubMedGoogle Scholar
  38. Eisenberg T, Glaeser SP, Ewers C, Semmler T, Drescher B, Kämpfer P. Caviibacter abscessus gen. nov., sp. nov., a member from the family Leptotrichiaceae isolated from guinea pigs (Cavia porcellus). Int J Syst Evol Microbiol. 2016;66(4):1652–9.View ArticlePubMedGoogle Scholar
  39. Kimura M, Tanikawa T, Suzuki M, Koizumi N, Kamiyama T, Imaoka K, Yamada A. Detection of Streptobacillus spp. in feral rats by specific polymerase chain reaction. Microbiol Immunol. 2008;52(1):9–15.View ArticlePubMedGoogle Scholar
  40. Rohde J, Rapsch C, Fehr M. Case report: Abscessation due to Streptobacillus moniliformis in a rat [in German]. Prakt Tierarzt. 2008;89(6):466–73.Google Scholar
  41. Eisenberg T, Ewers C, Rau J, Akimkin V, Nicklas W. Approved and novel strategies in diagnostics of rat bite fever and other Streptobacillus infections in humans and animals. Virulence. 2016;7(6):630–48.View ArticlePubMedGoogle Scholar
  42. Aziz RK, Bartels D, Best AA, DeJongh M, Disz T, Edwards RA, Formsma K, Gerdes S, Glass EM, Kubal M, et al. The RAST Server: rapid annotations using subsystems technology. BMC Genomics. 2008;9:75.View ArticlePubMedPubMed CentralGoogle Scholar
  43. Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, Buxton S, Cooper A, Markowitz S, Duran C, et al. Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics. 2012;28(12):1647–9.View ArticlePubMedPubMed CentralGoogle Scholar
  44. von Mentzer A, Connor TR, Wieler LH, Semmler T, Iguchi A, Thomson NR, Rasko DA, Joffre E, Corander J, Pickard D, et al. Identification of enterotoxigenic Escherichia coli (ETEC) clades with long-term global distribution. Nat Genet. 2014;46(12):1321–6.View ArticleGoogle Scholar
  45. Eisenberg T, Fawzy A, Nicklas W, Semmler T, Ewers C. Data from: Phylogenetic and comparative genomics of the family Leptotrichiaceae and introduction of a novel fingerprinting MLVA for Streptobacillus moniliformis. Dryad Digital Repository. doi:10.5061/dryad.1q7q4; 2016.
  46. Stamatakis A. RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics. 2014;30(9):1312–3.View ArticlePubMedPubMed CentralGoogle Scholar
  47. Hyatt D, Chen GL, Locascio PF, Land ML, Larimer FW, Hauser LJ. Prodigal: prokaryotic gene recognition and translation initiation site identification. BMC Bioinformatics. 2010;11:119.View ArticlePubMedPubMed CentralGoogle Scholar
  48. Marchler-Bauer A, Derbyshire MK, Gonzales NR, Lu S, Chitsaz F, Geer LY, Geer RC, He J, Gwadz M, Hurwitz DI, et al. CDD: NCBI’s conserved domain database. Nucleic Acids Res. 2015;43(Database issue):D222–6.View ArticlePubMedGoogle Scholar
  49. Legendre M, Pochet N, Pak T, Verstrepen KJ. Sequence-based estimation of minisatellite and microsatellite repeat variability. Genome Res. 2007;17(12):1787–96.View ArticlePubMedPubMed CentralGoogle Scholar
  50. Zhang Z, Schwartz S, Wagner L, Miller W. A greedy algorithm for aligning DNA sequences. J Comput Biol. 2000;7(1–2):203–14.View ArticlePubMedGoogle Scholar
  51. Yamamoto R, Clark GT. Streptobacillus moniliformis infection in turkeys. Vet Rec. 1966;79(4):95–100.View ArticlePubMedGoogle Scholar
  52. Wullenweber M, Kaspareit-Rittinghausen J, Farouq M. Streptobacillus moniliformis epizootic in barrier-maintained C57BL/6 J mice and susceptibility to infection of different strains of mice. Lab Anim Sci. 1990;40(6):608–12.PubMedGoogle Scholar
  53. Boot R, Oosterhuis A, Thuis HC. PCR for the detection of Streptobacillus moniliformis. Lab Anim. 2002;36(2):200–8.View ArticlePubMedGoogle Scholar
  54. Wullenweber M, Jonas C, Kunstyr I. Streptobacillus moniliformis isolated from otitis media of conventionally kept laboratory rats. J Exp Anim Sci. 1992;35(1):49–57.PubMedGoogle Scholar
  55. Kondruweit M, Weyand M, Mahmoud FO, Geissdorfer W, Schoerner C, Ropers D, Achenbach S, Strecker T. Fulminant endocarditis caused by Streptobacillus moniliformis in a young man. J Thorac Cardiovasc Surg. 2007;134(6):1579–80.View ArticlePubMedGoogle Scholar
  56. Loridant S, Jaffar-Bandjee MC, La Scola B. Shell vial cell culture as a tool for Streptobacillus moniliformis “resuscitation”. Am J Trop Med Hyg. 2011;84(2):306–7.View ArticlePubMedPubMed CentralGoogle Scholar
  57. Hopkinson WI, Lloyd JM. Streptobacillus moniliformis septicaemia in spinifex hopping mice (Notomys alexis). Aust Vet J. 1981;57(11):533–4.View ArticlePubMedGoogle Scholar
  58. Hanff PA, Rosol-Donoghue JA, Spiegel CA, Wilson KH, Moore LH. Leptotrichia sanguinegens sp. nov., a new agent of postpartum and neonatal bacteremia. Clin Infect Dis. 1995;20 Suppl 2:S237–9.View ArticlePubMedGoogle Scholar
  59. Collins MD, Hoyles L, Tornqvist E, von Essen R, Falsen E. Characterization of some strains from human clinical sources which resemble “Leptotrichia sanguinegens”: description of Sneathia sanguinegens sp. nov., gen. nov. Syst Appl Microbiol. 2001;24(3):358–61.View ArticlePubMedGoogle Scholar
  60. Sebald M. Etude sur les bacteries anaerobies gram-negatives asporulees. Laval: Theses de L’universite Paris, Imprimerie Barneoud S. A; 1962.Google Scholar
  61. Maher M, Palmer R, Gannon F, Smith TJ. Relationship of a novel bacterial fish pathogen to Streptobacillus moniliformis and the Fusobacteria group, based on 16S ribosomal RNA analysis. Syst Appl Microbiol. 1995;18:79–84.View ArticleGoogle Scholar
  62. Kapatral V, Anderson I, Ivanova N, Reznik G, Los T, Lykidis A, Bhattacharyya A, Bartman A, Gardner W, Grechkin G, et al. Genome sequence and analysis of the oral bacterium Fusobacterium nucleatum strain ATCC 25586. J Bacteriol. 2002;184(7):2005–18.View ArticlePubMedPubMed CentralGoogle Scholar

Copyright

© The Author(s). 2016

Advertisement