Open Access

Draft genome sequences and description of Lactobacillus rhamnosus strains L31, L34, and L35

  • Prapaporn Boonma
  • , Jennifer K. Spinler,
  • , Xiang Qin
  • , Chutima Jittaprasatsin
  • , Donna M. Muzny
  • , Harsha Doddapaneni
  • , Richard Gibbs
  • , Joe Petrosino,
  • , Somying Tumwasorn
  • and James Versalovic, ,
Corresponding author

DOI: 10.4056/sigs.5048907

Received: 15 February 2014

Accepted: 15 February 2014

Published: 15 June 2014

Abstract

Lactobacillus rhamnosus is a facultative, lactic acid bacterium in the phylum Firmicutes. Lactobacillus spp. are generally considered beneficial, and specific strains of L. rhamnosus are validated probiotics. We describe the draft genomes of three L. rhamnosus strains (L31, L34, and L35) isolated from the feces of Thai breastfed infants, which exhibit anti-inflammatory properties in vitro. The three genomes range between 2.8 – 2.9 Mb, and contain approximately 2,700 protein coding genes.

Keywords:

Lactobacillus rhamnosuscomparative genomicsprobioticslactic acid bacteriaanti-inflammatory

Introduction

Lactobacillus is the largest of three genera within the family Lactobacillaceae, and belongs to one of the dominant phyla, Firmicutes, in the human microbiome [1]. Lactobacillus spp. are naturally isolated from fermented foods [2], and are key members of the human microbiota, reviewed in [3]. In humans, they colonize the oral cavity, gastrointestinal and urogenital tracts, and breast milk [4]. As a whole, this genus is beneficial to humans, possesses many probiotic traits, and is rarely associated with disease.

The human-intestinal isolate, L. rhamnosus strain GG, is one of the most studied and applied probiotics. Research has shown that L. rhamnosus GG can modulate host immunity in vitro by decreasing inflammatory cytokine production from various eukaryotic cell lines [5,6], induces intestinal mucin gene expression subsequently inhibiting pathogen adherence in vitro [7]; and attenuates in vitro barrier dysfunction induced by inflammatory cytokines [8]. Here we present the draft genomes and classification summary of three potential probiotic L. rhamnosus strains L31, L34, and L35 isolated from the feces of Thai breastfed infants [9]. Genome sequencing and comparisons of L31, L34, and L35 with the species type-strain, L. rhamnosus GG should help researchers identify distinguishing genetic features important for specific probiotic traits.

Classification and features

Within the phylum Firmicutes, the family Lactobacillaceae contains three genera: Lactobacillus, Paralactobacillus, and Pediococcus; Lactobacillus being the largest with latest estimates ranging between 227-230 species (Web Site) [10]. Members of Lactobacillus are gram-positive, non-motile, anaerobic, lactic-acid-producing bacilli that are divided into three fermentation groups: A) obligately homofermentative, B) facultatively heterofermentative, and C) obligately heterofermentative [4]. L. rhamnosus resides in fermentation group B and is distinct from the three major Lactobacillus phylogenetic groups based on 16S rRNA gene sequence (L. delbrueckii, L. reuteri, and L. salivarius groups) [4]. L. rhamnosus strains L31, L34, and L35 are phylogenetically similar to L. rhamnosus GG and maintain a distinctive 16S rRNA gene-based phyologeny from the three major Lactobacillus groups (Figure 1). The basic characteristics of L. rhamnosus L31, L34, and L35 are summarized in Table 1.

Figure 1

The phylogenetic tree represents the relationships of L. rhamnosus strains L31, L34, and L35 with respect to several members of the genus Lactobacillus. The strains and their corresponding GenBank accession numbers for 16S rRNA genes are: L. rhamnsosus strain GG, NC_013198, L. salivarius strain CECT 5713, NC_017481, L. ruminis strain ATCC 27782, NC_015975, L. reuteri strain JCM 1112, NC_010609, L. fermentum strain CECT 5716, NC_017465, L. johnsonii strain NCC 533, NC_005362, L. delbrueckii subsp. bulgaricus strain ATCC 11842, NC_008054, L. acidophilus strain NCFM, NC_006814. Full-length 16S rRNA gene sequences were aligned using ClustalW, and phylogenetic inferences were obtained using the maximum-likelihood method within the MEGA 5.2 software [11] with 1,000 bootstraps. B. subtilis strain 6051 HGW (NC_020507) was used as an outgroup.

Table 1

Classification and general features of L. rhamnosus strains L31, L34, and L35 according to the MIGS recommendation

MIGS ID

        Property

        Term

       Evidence codea

        Classification

        Domain Bacteria

       TAS [12]

        Phylum Firmicutes

       TAS [13-15]

        Class Bacillus

       TAS [16-18]

        Order Lactobacillales

       TAS [19,20]

        Family Lactobacillaceae

       TAS [16,21]

        Genus Lactobacillus

       TAS [16,22-26]

        Species Lactobacillus rhamnosus

       TAS [27]

        Strains L31, L34, and L35

       IDA

        Gram stain

        Positive

       IDA

        Cell shape

        Rod-shaped

       IDA

        Motility

        Non-motile

       NAS

        Sporulation

        Non-sporulating

       NAS

        Temperature range

        Mesophile

       NAS

        Optimum temperature

        37°C

       IDA

        Carbon source

        Glucose

       NAS

        Energy source

        Lactose, glucose and other sugars

       NAS

MIGS-6

        Habitat

        Human GI Tract

       NAS

MIGS-22

        Oxygen

        Facultative anaerobes

       IDA

MIGS-15

        Biotic relationship

        Symbiotic relationship

       NAS

MIGS-14

        Pathogenicity

        Nonpathogenic; potential probiotic

       IDA

        Biosafety level

        1

       NAS

        Isolation

        Infant Feces

       IDA

MIGS-4

        Geographic location

        Bangkok, Thailand

       IDA

MIGS-5

        Sample collection time

        Not reported

MIGS-4.1

        Latitude

        13° 45’N

       IDA

MIGS-4.2

        Longitude

        100° 35’E

       IDA

MIGS-4.4

        Altitude

        Not reported

       NAS

a) Evidence codes – IDA: Inferred from Direct Assay; 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). These evidence codes are from the Gene Ontology project [28].

The colony and Gram stain morphology of L. rhamnosus strains L31, L34, and L35 are each depicted in Figure 2. Supernatants from L. rhamnosus L34 and L35, both isolated from the same 40 day old female, suppress LPS-induced TNF-α production by THP-1 cells [9] and C. difficile-induced IL-8 production by HT-29 cells [29]. Similarly, strain L31, isolated from a 39 day old female, suppresses LPS-induced TNF-α production by THP-1 cells, however does not suppress C. difficile-induced IL-8 production by HT-29 cells [29]. All three strains are resistant to two drugs commonly used to treat C. difficile infection in humans, vancomycin and metronidazole (MIC90 >256µg/mL for each), but are susceptible to low concentrations (MIC90 = 2µg/mL) of the newest antibiotic targeting C. difficile, fidaxomicin. These strain-specific characteristics suggest L. rhamnosus L34 and L35 are potential probiotic candidates for either preventing or treating C. difficile disease.

Figure 2

Colony morphology and Gram stains of L. rhamnosus strains L31, L34, and L35. L. rhamnosus strains were cultured anaerobically on MRS agar at 37°C for 48 hr. Gram stains were carried out using standard methods, and images were taken under oil emersion at 100× magnification.

Genome sequencing information

Genome project history

L. rhamnosus strains L31, L34, and L35 were selected for sequencing based on the properties described above. The draft genome sequence for each strain was finished in October 2012. The Whole Genome Shotgun projects for L. rhamnosus L31, L34, and L35 have been deposited at DDBJ/EMBL/GenBank under the accession numbers AYTQ00000000, AYTR00000000, and AYTP0000000, respectively. The versions described in this paper are AYTQ01000000, AYTR01000000, and AYTP0100000, respectively. The genome projects for L31, L34, and L35 are listed in the Genome OnLine Database (GOLD) [30] as projects Gi0036900, Gi0036903, and Gi0036905, respectively. Genome sequencing and assembly was completed at Baylor College of Medicine’s Human Genome Sequencing Center (BCM-HGSC). Automatic annotation was performed using the DOE-JGI Microbial Annotation Pipeline (DOE-JGI MAP). Table 2 shows the project information and its association with MIGS version 2.0 compliance [31].

Table 2

Project information

MIGS ID

      L31

      L34

       L35

MIGS ID

      Property

      Term

      Term

       Term

MIGS-31

      Finishing quality

      Standard Draft

      Standard Draft

       Standard Draft

MIGS-28

      Libraries used

      8 kb, mate paired library

      8 kb, mate paired library

       8 kb, mate paired library

MIGS-29

      Sequencing platforms

      454 GS FLX

      454 GS FLX

       454 GS FLX

MIGS-31.2

      Fold coverage

      23×

      29×

       26×

MIGS-30

      Assemblers

      Newbler v2.5.3

      Newbler v2.5.3

       Newbler v2.5.3

MIGS-32

      Gene calling method

      Prodigal

      Prodigal

       Prodigal

      Genome Database release

      March 1, 2014

      March 1, 2014

       March 1, 2014

      GenBank ID

      AYTQ00000000

      AYTR00000000

       AYTP00000000

      GenBank Date of Release

      March 1, 2014

      March 1, 2014

       March 1, 2014

      GOLD ID

      Gi0036900

      Gi0036903

       Gi0036905

      Project relevance

      Potential probiotic

      Potential probiotic

       Potential probiotic

Growth conditions and DNA isolation

L. rhamnosus strains L31, L34, and L35 were routinely cultured in an anaerobic chamber (Concept Plus, Ruskinn Technology, UK) (10% CO2, 10% H2, and 80% N2) for 24-48 h at 37°C in de Man, Rogosa, Sharpe (MRS) medium (Oxoid, England). For genomic DNA isolation, cultures were adjusted to an OD600 of 0.1 and incubated anaerobically at 37°C for 8 h. Bacterial pellets were collected by centrifugation and the DNA was extracted using QIAGEN Genomic-tip100/G columns (Qiagen, Germany) according to the manufacturer’s instructions. DNA quality was analyzed by agarose gel electrophoresis, and concentrations were determined by fluorescence using the Qubit™ DNA Assay (Life Technologies, USA).

Genome sequencing and assembly

The genomes of L. rhamnosus strains L31, L34, and L35 were sequenced at the BCM-HGSC, USA on a Roche 454 GS FLX sequencing platform. A fragment sequencing approach was implemented using 8 kb libraries generated by long insert mate paired construction, as detailed in the Human Microbiome Project Reference Genome Project protocol [32] to about 23× (254,342 reads), 29× (283,036 reads), and 26× (249,176 reads) sequence depth coverage, respectively, with an estimated read alignment error rate of 0.84%. The sequence data were assembled using the Newbler assembler version 2.5.3. The final assemblies resulted in 67 (L31), 51 (L34), and 51 (L35) contigs generating corresponding genome sizes of 2.8, 2.9, and 2.9 Mb in 3, 3, and 4 scaffolds.

Genome annotation

Open Reading Frames (ORFs) were predicted using Prodigal [33,34] as part of the Oak Ridge National Laboratory genome annotation pipeline, followed by a round of manual curation using the JGI GenePRIMP pipeline [35]. The predicted protein coding sequences (CDSs) were translated and searched against the National Center for Biotechnology Information (NCBI) non-redundant database, UniProt, TIGRFam, Pfam, PRIAM, KEGG, COG, and InterPro databases [35]. These data sources were combined to assert a product description for each predicted protein. Additional gene prediction analysis and manual functional annotation was performed with the Integrated Microbial Genomes Expert Review (IMG-ER) platform [36]. Non-coding genes and miscellaneous features were predicted using tRNAscan-SE [37], RNAMMer [38], Rfam [39], TMHMM [40], and signalP [41].

Genome properties

The properties and statistics for the three L. rhamnosus genomes are summarized in Table 3. The distribution of genes into COG functional categories for each genome is detailed in Table 4. The L. rhamnosus L31 genome was assembled into 67 contigs (ranging from 551 – 290,053 bp) forming one presumptive circular chromosome of 2,826,754 base pairs (46.73% GC content). A total of 2,749 ORFs were predicted: 2,687 are protein-coding genes, and 62 are RNA genes. A total of 2,173 (79.05%) protein-coding genes were assigned a putative function. The L34 genome was assembled into 51 contigs (ranging from 288 – 237,520 bp) forming a presumptive single circular chromosome of 2,937,717 base pairs (46.81% GC content). A total of 2,845 ORFs were predicted: 2,774 are protein-coding genes, and 71 are RNA genes. A total of 2,216 (77.89%) protein coding genes were assigned a putative function. Finally, the L35 genome was assembled into 51 contigs (687 – 226,797 bp) forming one presumptive chromosome of 2,937,403 base pairs (46.81%). A total of 2,842 ORFs were predicted: 2,772 are protein-coding genes, and 70 are RNA genes. A total of 2,217 (78.01%) protein coding genes were assigned a putative function.

Table 3

Nucleotide content and gene count level of the genomes

      L31

     L34

     L35

Attribute

     Value

      %agea

     Value

     %agea

     Value

     %agea

Genome Size (bp)

     2,826,754

      100

     2,937,717

     100

     2,937,403

     100

DNA G+C content (bp)

     1,320,949

      46.73

     1,375,266

     46.81

     1,375,134

     46.81

DNA coding region (bp)

     2,422,731

      85.71

     2,519,202

     85.75

     2,517,453

     85.70

Total genes

     2,749

      100

     2,854

     100

     2,842

     100

RNA genes

     62

      2.26

     71

     2.50

     70

     2.46

Protein-coding genes

     2,687

      97.74

     2,774

     97.50

     2,772

     97.54

Genes with functional prediction

     2,173

      79.05

     2,216

     77.89

     2,217

     78.01

Genes in paralog clusters

     1,818

      66.13

     1,898

     66.71

     1,869

     65.76

Genes assigned to COGs

     2,121

      77.16

     2,150

     75.57

     2,151

     75.69

Genes assigned to KOGs

     886

      32.23

     913

     32.09

     914

     32.16

Genes assigned to Pfam

     2,209

      80.36

     2,250

     79.09

     2,254

     79.31

Genes assigned to TIGRfam

     880

      31.01

     893

     31.39

     892

     31.39

Genes with signal peptides

     138

      5.02

     139

     4.89

     138

     4.86

Genes with transmembrane helices

     813

      29.57

     835

     29.35

     834

     29.35

a) The total is based on either the size of the genome in base pairs or the total number of protein coding genes in the annotated genome.

Table 4

Number of genes associated with the 25 general COG functional categories

      L31

       L34

      L35

Code

      Value

      %agea

      Value

       %agea

       Value

      %agea

       Description

J

      150

      6.56

      150

       6.49

       150

      6.48

       Translation

A

      -

      -

      -

       -

       -

      -

       RNA processing and modification

K

      203

      8.88

      206

       8.91

       207

      8.95

       Transcription

L

      123

      5.38

      132

       5.71

       134

      5.79

       Replication, recombination and repair

B

      -

      -

      -

       -

       -

      -

       Chromatin structure and dynamics

D

      33

      1.44

      29

       1.25

       29

      1.25

       Cell cycle control, mitosis and meiosis

Y

      -

      -

      -

       -

       -

      -

       Nuclear structure

V

      75

      3.28

      79

       3.42

       79

      3.41

       Defense mechanisms

T

      82

      3.59

      84

       3.63

       85

      3.67

       Signal transduction mechanisms

M

      129

      5.65

      128

       5.54

       127

      5.49

       Cell wall/membrane biogenesis

N

      9

      0.39

      8

       0.35

       8

      0.35

       Cell motility

Z

      -

      -

      -

       -

       -

      -

       Cytoskeleton

W

      -

      -

      -

       -

       -

      -

       Extracellular structures

U

      28

      1.23

      23

       0.99

       23

      0.99

       Intracellular trafficking and secretion

O

      59

      2.58

      59

       2.55

       59

      2.5

       Posttranslational modification, protein turnover, chaperones

C

      90

      3.94

      87

       3.76

       88

      3.80

       Energy production and conversion

G

      303

      13.26

      315

       13.62

       315

      13.61

       Carbohydrate transport and metabolism

E

      183

      8.01

      178

       7.70

       177

      7.65

       Amino acid transport and metabolism

F

      87

      3.81

      85

       3.68

       85

      3.67

       Nucleotide transport and metabolism

H

      58

      2.54

      60

       2.60

       60

      2.59

       Coenzyme transport and metabolism

I

      55

      2.41

      57

       2.47

       57

      2.46

       Lipid transport and metabolism

P

      96

      4.20

      95

       4.11

       95

      4.11

       Inorganic ion transport and metabolism

Q

      21

      0.92

      21

       0.91

       21

      0.91

       Secondary metabolites biosynthesis, transport and catabolism

R

      285

      12.47

      292

       12.63

       291

      12.58

       General function prediction only

S

      216

      9.45

      224

       9.69

       224

      9.68

       Function unknown

-

      628

      22.84

      695

       24.43

       691

      24.31

       Not in COGs

a) The total is based on the total number of protein coding genes in the annotated genome.

Comparison with Lactobacillus rhamnosus strain GG

The beneficial effects of human-intestinal derived L. rhamnosus GG have been studied for two decades [42-45] and its complete genome is available in NCBI [46]. We have compared the draft genome sequences of the potential probiotic L. rhamnosus strains L31, L34, and L35 to L. rhamnosus GG. The L. rhamnosus GG genome (3,010,111 bp, 46.69% GC content) is slightly larger than the new genomes presented here, and has approximately the same GC content (Table 3). In a recent comparative genomics study of 100 L. rhamnosus strains, Douillard, et al. [47] delineated seventeen variable chromosomal regions of L. rhamnosus strain GG (annotated in Figure 3), and the majority of these regions are absent or incomplete in the genomes of strains L31, L34, and L35 (Figure 3), notably the spaCBA pili gene cluster required for mucus adhesion [46]. The galactitol PTS region important for dulcitol utilization, a trait that typically belongs to L. rhamnosus isolates adapted to the intestinal tract [47], is conserved in L31, L34, and L35. Similar to L. rhamnosus GG, L31, L34, and L35 each contain genes annotated as L-lactate dehydrogenase (ldhL) and D-lactate dehydrogenase (ldhD) important for synthesizing L-lactate and D-lactate from pyruvate, respectively [49]. L. rhamnosus GG is unable to metabolize either maltose due to an inserted gene between the maltose-specific transport genes and hydrolase, or lactose because of a 38 bp N-terminal truncation in lacT and a disrupted lacG [47,50]. Strains L31, L34, and L35 all have an intact maltose locus and carry non-mutated copies of lacT and lacG (locations indicated on Figure 3), and therefore are predicted to utilize both maltose and lactose.

Figure 3

Circular representation of 3 draft L. rhamnosus genomes compared against L. rhamnosus strain GG (NC_ 013198). The innermost rings show GC content (black) and GC skew (purple/green). The remaining rings show BLASTn results of each genome against L. rhamnosus GG with results rendered using the BRIG program [48]. Relative shading density (from darker to lighter) within each circle represents levels of nucleotide homology. Blank regions represent absent genetic regions. Genetic regions of interest are annotated on the outermost ring. Numbered elements (1-17) represent the previously identified variable chromosomal regions of L. rhamnosus GG [47].

In line with the anti-inflammatory phenotypic differences already noted [9,29], differences in genomic features between L. rhamnosus L31 and the two isolates, L34 and L35, can also be made relative to strain GG. The taurine transport system deemed important for bile resistance as well as the fucU, fucI, fcsR, and α-L-fucosidase genes required for metabolizing fucosylated compounds present in gastrointestinal environments are found in L34 and L35 genomes, but not in L31. L. rhamnosus GG, despite belonging to a species known for rhamnose utilization, possesses an altered rhamnose locus and cannot utilize rhamnose [46]. L. rhamnosus L31 contains an intact rhamnose locus, while this locus in strains L34 and L35 looks similarly disrupted to that of strain GG. It is also noteworthy that L. rhamnosus L31 contains an iron-transport and a general secretion system not present in strains L34, L35, or GG.

Conclusion

Here we have presented the draft genomes of three potential probiotic strains of L. rhamnosus: L31, L34, and L35. Brief genome comparisons indicate that strains L34 and L35 are most similar to L. rhamnosus GG, while L31 contains marked differences suggesting it may have originated from a slightly different ecological niche [47]. L. rhamnosus L34 and L35 were isolated from the same host based on initial distinguishing colony morphology [9], however current colony morphology for these strains is not unique (Figure 2) and comparison of the draft genomes suggests the two genomes are nearly identical and similarly distinct from L31. It is possible that L34 and L35 may represent isolates of the same strain. Future studies will combine functional data with genomics, which is a powerful method for not only validating probiotic features of beneficial microbes, but also for learning about the environmental adaptations that have favored their mutual relationship with human hosts.

Declarations

Acknowledgements

The authors would like to acknowledge the Texas Children’s Microbiome Center for providing equipment and resources for a fruitful collaboration. This work was supported by the NIH/National Institute of Diabetes and Digestive and Kidney Disease Grants P30 DK56338 (JV) and 5UH3DK083990-04 (JV), as well as the Thailand Research Fund through the Royal Golden Jubilee PhD Program (PHD/0295/2550) (PB), and the Rachadapisek Sompoj Research Fund, Faculty Medicine, Chulalongkorn University (Grant No. RA51/1 and RA55/20) (ST).


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References

  1. . Structure, function and diversity of the healthy human microbiome. Nature. 2012; 486:207-214 View ArticlePubMed
  2. Bernardeau M, Vernoux JP, Henri-Dubernet S and Guéguen M. Safety assessment of dairy microorganisms: the Lactobacillus genus. Int J Food Microbiol. 2008; 126:278-285 View ArticlePubMed
  3. Spinler JK. Human Microbiome, Lactobacillaceae in the. In: Nelson K, editor. Encyclopedia of Metagenomics. Verlag Berlin Heidelberg: Springer; 2013.
  4. Schleifer KH. Family I. Lactobacillaceae Winslow, Broadhurst, Buchanan, Krumwiede, Rogers and Smith 1917, familia. In: De Vos PG, G.M.; Jones, D.; Krieg, N.R.; Ludwig, W.; Rainey, F.A.; Schleifer, K.H.; Whitman, W.B., editor. Bergey's Manual of Systematic Bacteriology, Volume Three The Firmicutes 2nd ed. Volume 3, The Firmicutes. New York: Springer; 2009. p 465-532.
  5. Peña JA and Versalovic J. Lactobacillus rhamnosus GG decreases TNF-alpha production in lipopolysaccharide-activated murine macrophages by a contact-independent mechanism. Cell Microbiol. 2003; 5:277-285 View ArticlePubMed
  6. Zhang L, Li N, Caicedo R and Neu J. Alive and dead Lactobacillus rhamnosus GG decrease tumor necrosis factor-alpha-induced interleukin-8 production in Caco-2 cells. J Nutr. 2005; 135:1752-1756PubMed
  7. Mack DR, Michail S, Wei S, McDougall L and Hollingsworth MA. Probiotics inhibit enteropathogenic E. coli adherence in vitro by inducing intestinal mucin gene expression. Am J Physiol. 1999; 276:G941-G950PubMed
  8. Donato KA, Gareau MG, Wang YJ and Sherman PM. Lactobacillus rhamnosus GG attenuates interferon-{gamma} and tumour necrosis factor-alpha-induced barrier dysfunction and pro-inflammatory signalling. Microbiology. 2010; 156:3288-3297 View ArticlePubMed
  9. Jittaprasatsin C. Quantification and determination of antagonistic activity of bifidobacteria and lactobacilli in faeces of breast-fed and mixed-fed infants. Bangkok,Thailand: Chulalongkorn University; 2008. 113 p.
  10. Euzéby JP. List of Bacterial Names with Standing in Nomenclature: a folder available on the Internet. Int J Syst Bacteriol. 1997; 47:590-592 View ArticlePubMed
  11. Tamura K, Peterson D, Peterson N, Stecher G, Nei M and Kumar S. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol. 2011; 28:2731-2739 View ArticlePubMed
  12. Woese CR, Kandler O and Wheelis ML. Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc Natl Acad Sci USA. 1990; 87:4576-4579 View ArticlePubMed
  13. Gibbons NE and Murray RGE. Proposals concerning the higher taxa of bacteria. Int J Syst Bacteriol. 1978; 28:1-6 View Article
  14. Garrity G, Holt J. The Road Map to the Manual. Bergey's Manual of Systematic Bacteriology. Volume 1. New York: Springer; 2001. p 119-169.
  15. Murray RGE. The Higher Taxa, or, a Place for Everything...? In: Holt JG (ed), Bergey's Manual of Systematic Bacteriology, First Edition, Volume 1, The Williams and Wilkins Co., Baltimore, 1984, p. 31-34.
  16. Skerman VBD, McGowan V and Sneath PHA. Approved Lists of Bacterial Names. Int J Syst Bacteriol. 1980; 30:225-420 View Article
  17. Cohn F. Untersuchungen über Bakterien. Beitr Biol Pflanz. 1872; 1:127-224
  18. Gibson T, Gordon RE. Genus I. Bacillus Cohn 1872, 174; Nom. gen. cons. Nomencl. Comm. Intern. Soc. Microbiol. 1937, 28; Opin. A. Jud. Comm. 1955, 39. In: Buchanan RE, Gibbons NE (eds), Bergey's Manual of Determinative Bacteriology, Eighth Edition, The Williams and Wilkins Co., Baltimore, 1974, p. 529-550.
  19. List of new names and new combinations previously effectively, but not validly, published. List no. 132. Int J Syst Evol Microbiol. 2010; 60:469-472 View Article
  20. Ludwig W, Schleifer KH, Whitman WB. Order II. Lactobacillales ord. nov. In: De Vos P, Garrity G, Jones D, Krieg NR, Ludwig W, Rainey FA, Schleifer KH, Whitman WB (eds), Bergey's Manual of Systematic Bacteriology, Second Edition, Volume 3, Springer-Verlag, New York, 2009, p. 464.
  21. Winslow CEA, Broadhurst J, Buchanan RE, Krumwiede C, Rogers LA and Smith GH. The Families and Genera of the Bacteria: Preliminary Report of the Committee of the Society of American Bacteriologists on Characterization and Classification of Bacterial Types. J Bacteriol. 1917; 2:505-566PubMed
  22. Beijerinck MW. Sur les ferments lactiques de l'industrie. Archives Néerlandaises des Sciences Exactes et Naturelles. 1901; 6:212-243
  23. Cai Y, Pang H, Kitahara M and Ohkuma M. Lactobacillus nasuensis sp. nov., a lactic acid bacterium isolated from silage, and emended description of the genus Lactobacillus. Int J Syst Evol Microbiol. 2012; 62:1140-1144 View ArticlePubMed
  24. Rogosa M. Genus Lactobacillus Beijerinck 1901, 212; Nom. cons. Opin. 38, Jud. Comm. 1971, 104. In: Buchanan RE, Gibbons NE (eds), Bergey's Manual of Determinative Bacteriology, Eighth Edition, The Williams and Wilkins Co., Baltimore, 1974, p. 576-593.
  25. Haakensen M, Dobson CM, Hill JE and Ziola B. Reclassification of Pediococcus dextrinicus (Coster and White 1964) Back 1978 (Approved Lists 1980) as Lactobacillus dextrinicus comb. nov., and emended description of the genus Lactobacillus. Int J Syst Evol Microbiol. 2009; 59:615-621 View ArticlePubMed
  26. Editorial Secretary (for the Judicial Commission of the International Committee on Systematic Bacteriology). Opinion 38: Conservation of the Generic Name Lactobacillus Beijerinck. Int J Syst Bacteriol. 1971; 21:104 View Article
  27. Felis GE and Dellaglio F. Taxonomy of Lactobacilli and Bifidobacteria. Curr Issues Intest Microbiol. 2007; 8:44-61PubMed
  28. Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, Davis AP, Dolinski K, Dwight SS and Eppig JT. Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat Genet. 2000; 25:25-29 View ArticlePubMed
  29. Boonma P. Role of Lactobacillus in the suppression of Clostridium difficile-induced IL-8 production in colonic epithelial cells. Bangkok,Thailand: Chulalongkorn University; 2013. 120 p.
  30. Liolios K, Chen IM, Mavromatis K, Tavernarakis N, Hugenholtz P, Markowitz VM and Kyrpides NC. The Genomes On Line Database (GOLD) in 2009: status of genomic and metagenomic projects and their associated metadata. Nucleic Acids Res. 2010; 38:D346-D354 View ArticlePubMed
  31. Field D, Garrity G, Gray T, Morrison N, Selengut J, Sterk P, Tatusova T, Thomson N, Allen MJ and Angiuoli SV. The minimum information about a genome sequence (MIGS) specification. Nat Biotechnol. 2008; 26:541-547 View ArticlePubMed
  32. Nelson KE, Weinstock GM, Highlander SK, Worley KC, Creasy HH, Wortman JR, Rusch DB, Mitreva M, Sodergren E and Chinwalla AT. A catalog of reference genomes from the human microbiome. Science. 2010; 328:994-999 View ArticlePubMed
  33. Claesson MJ, van Sinderen D and O'Toole PW. Lactobacillus phylogenomics--towards a reclassification of the genus. Int J Syst Evol Microbiol. 2008; 58:2945-2954 View ArticlePubMed
  34. Hyatt D, Chen GL, Locascio PF, Land ML, Larimer FW and Hauser LJ. Prodigal: prokaryotic gene recognition and translation initiation site identification. BMC Bioinformatics. 2010; 11:119 View ArticlePubMed
  35. Pati A, Ivanova NN, Mikhailova N, Ovchinnikova G, Hooper SD, Lykidis A and Kyrpides NC. GenePRIMP: a gene prediction improvement pipeline for prokaryotic genomes. Nat Methods. 2010; 7:455-457 View ArticlePubMed
  36. Markowitz VM, Mavromatis K, Ivanova NN, Chen IM, Chu K and Kyrpides NC. IMG ER: a system for microbial genome annotation expert review and curation. Bioinformatics. 2009; 25:2271-2278 View ArticlePubMed
  37. Lowe TM and Eddy SR. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res. 1997; 25:955-964 View ArticlePubMed
  38. Lagesen K, Hallin P, Rodland EA, Staerfeldt HH, Rognes T and Ussery DW. RNAmmer: consistent and rapid annotation of ribosomal RNA genes. Nucleic Acids Res. 2007; 35:3100-3108 View ArticlePubMed
  39. Griffiths-Jones S, Bateman A, Marshall M, Khanna A and Eddy SR. Rfam: an RNA family database. Nucleic Acids Res. 2003; 31:439-441 View ArticlePubMed
  40. Krogh A, Larsson B, von Heijne G and Sonnhammer EL. Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. J Mol Biol. 2001; 305:567-580 View ArticlePubMed
  41. Bendtsen JD, Nielsen H, von Heijne G and Brunak S. Improved prediction of signal peptides: SignalP 3.0. J Mol Biol. 2004; 340:783-795 View ArticlePubMed
  42. Saxelin M, Pessi T and Salminen S. Fecal recovery following oral administration of Lactobacillus strain GG (ATCC 53103) in gelatine capsules to healthy volunteers. Int J Food Microbiol. 1995; 25:199-203 View ArticlePubMed
  43. Guarino A, Lo Vecchio A and Canani RB. Probiotics as prevention and treatment for diarrhea. Curr Opin Gastroenterol. 2009; 25:18-23 View ArticlePubMed
  44. Saxelin M, Tynkkynen S, Mattila-Sandholm T and de Vos WM. Probiotic and other functional microbes: from markets to mechanisms. Curr Opin Biotechnol. 2005; 16:204-211 View ArticlePubMed
  45. Vanderhoof JA and Mitmesser SH. Probiotics in the management of children with allergy and other disorders of intestinal inflammation. Benef Microbes. 2010; 1:351-356 View ArticlePubMed
  46. Kankainen M, Paulin L, Tynkkynen S, von Ossowski I, Reunanen J, Partanen P, Satokari R, Vesterlund S, Hendrickx AP and Lebeer S. Comparative genomic analysis of Lactobacillus rhamnosus GG reveals pili containing a human- mucus binding protein. Proc Natl Acad Sci USA. 2009; 106:17193-17198 View ArticlePubMed
  47. Douillard FP, Ribbera A, Kant R, Pietila TE, Jarvinen HM, Messing M, Randazzo CL, Paulin L, Laine P and Ritari J. Comparative Genomic and Functional Analysis of 100 Lactobacillus rhamnosus Strains and Their Comparison with Strain GG. PLoS Genet. 2013; 9:e1003683 View ArticlePubMed
  48. Alikhan NF, Petty NK, Ben Zakour NL and Beatson SA. BLAST Ring Image Generator (BRIG): simple prokaryote genome comparisons. BMC Genomics. 2011; 12:402 View ArticlePubMed
  49. Ferain T, Garmyn D, Bernard N, Hols P and Delcour J. Lactobacillus plantarum ldhL gene: overexpression and deletion. J Bacteriol. 1994; 176:596-601PubMed
  50. Tsai YK and Lin TH. Sequence, organization, transcription and regulation of lactose and galactose operons in Lactobacillus rhamnosus TCELL-1. J Appl Microbiol. 2006; 100:446-459 View ArticlePubMed