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Genome sequencing of Clostridium butyricum DKU-01, isolated from infant feces.

Mo S, Kim BS, Yun SJ, Lee JJ, Yoon SH, Oh CH - Gut Pathog (2015)

Bottom Line: The extracted 16S rRNA gene from genome sequences of DKU-01 was similar to Clostridium butyricum with 99.63% pairwise similarity.Genes related to Fructooligosaccharide utilization were detected in the genome of strain DKU-01 and compared with other genera, such as Bifidobacterium and Streptococcus.We found that strain DKU-01 can metabolize a wide range of carbohydrates in comparative genome result.

View Article: PubMed Central - PubMed

Affiliation: Biosafety & Validation Center, Clinical Trial Institute, Dankook University, Choenan, 330-714 Republic of Korea.

ABSTRACT

Background: Clostridium butyricum is a butyric acid-producing anaerobic bacteriuma, and commonly present as gut microbiota in humans. This species has been used as a probiotic for the prevention of diarrhea in humans. In this study, we report the draft genome of C. butyricum DKU-01, which was isolated from infant feces, to better understand the characteristics of this strain so that it can later be used in the development of probiotic products.

Results: A total of 79 contigs generated by hybrid assembly of sequences obtained from Roche 454 and Illumina Miseq sequencing systems were investigated. The assembled genome of strain DKU-01 consisted of 4,519,722 bp (28.62% G + C content) with a N50 contig length of 108,221 bp and 4,037 predicted CDSs. The extracted 16S rRNA gene from genome sequences of DKU-01 was similar to Clostridium butyricum with 99.63% pairwise similarity. The sequence of strain DKU-01 was compared with previously reported genome sequences of C. butyricum. The value of average nucleotide identity between strains DKU-01 and C. butyricum 60E3 was 98.74%, making it the most similar strain to DKU-01.

Conclusions: We sequenced the DKU-01 strain isolated from infant feces, and compared it with the available genomes of C. butyricum on a public database. Genes related to Fructooligosaccharide utilization were detected in the genome of strain DKU-01 and compared with other genera, such as Bifidobacterium and Streptococcus. We found that strain DKU-01 can metabolize a wide range of carbohydrates in comparative genome result. Further analyses of the comparative genome and fermentation study can provide the information necessary for the development of strain DKU-01 for probiotics.

No MeSH data available.


Related in: MedlinePlus

Comparison of themsmcluster in contig number 22 of DKU-01 genome with selected bacteria (based on RAST annotation server). The red arrow (number 1) indicates MsmE; yellow-green arrow (number 2) MsmF; brown arrow (number 3) MsmG; blue arrow (number 4) Oligo-1,6-glucosidase; yellow arrow (number 5) Alpha-galactosidase; blue-green (number 6) Catabolite control protein A; pink arrow (number 7) possible NagC/XylR-type transcriptional regulator; green arrow (number 8) tRNA-specific adenosine-34 deaminase; dark brown arrow (number 9) probable NhaP-type Na(+)/H(+) exchanger; sky-blue arrow (number 10) Sucrose phosphorylase; dark-gray arrow (number 11) MSM operon regulatory protein; pistachio arrow (number 12) Threonine dehydratase; lime-green arrow (number 13) Aldehyde dehydrogenase B; purple arrow (number 14) NAD-dependent protein deacetylase of SIR2 family; primrose arrow (number 15) possible phospholipase. The same number and color in the figure are sets of genes with similar sequences. Genes whose relative position is conserved in at least four other species are functionally coupled and shared gray background boxes.
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Fig3: Comparison of themsmcluster in contig number 22 of DKU-01 genome with selected bacteria (based on RAST annotation server). The red arrow (number 1) indicates MsmE; yellow-green arrow (number 2) MsmF; brown arrow (number 3) MsmG; blue arrow (number 4) Oligo-1,6-glucosidase; yellow arrow (number 5) Alpha-galactosidase; blue-green (number 6) Catabolite control protein A; pink arrow (number 7) possible NagC/XylR-type transcriptional regulator; green arrow (number 8) tRNA-specific adenosine-34 deaminase; dark brown arrow (number 9) probable NhaP-type Na(+)/H(+) exchanger; sky-blue arrow (number 10) Sucrose phosphorylase; dark-gray arrow (number 11) MSM operon regulatory protein; pistachio arrow (number 12) Threonine dehydratase; lime-green arrow (number 13) Aldehyde dehydrogenase B; purple arrow (number 14) NAD-dependent protein deacetylase of SIR2 family; primrose arrow (number 15) possible phospholipase. The same number and color in the figure are sets of genes with similar sequences. Genes whose relative position is conserved in at least four other species are functionally coupled and shared gray background boxes.

Mentions: We focused on fructooligosaccharide (FOS) utilization within the carbohydrate subsystem, because fructooligosaccharide has been used as a prebiotic in food products and infant formulas [20]. FOS can be a derivative of simple fructose polymers of fructose moieties attached to a sucrose molecule, and it can be degraded by a variety of lactic acid bacteria in the gut [21,22]. Lactobacillus acidophilus and Bifidobacterium spp. have been studied for the utilization of FOS [22,23], and the utilization of FOS by Clostridium spp. has also been reported [24]. Ten ORFs in the genome of strain DKU-01 were annotated to genes related to FOS utilization (Table 1). The chromosomal region including MsmE, MsmF, and MsmG in contig number 22 of strain DKU-01 was compared with synteny in C. botulinum E3, C. beijerincki NCIMB 8052, Bifidobacterium longum NCC 2705, B. animalis subsp. lactis AD011, B. adolescentis ATCC 15703, Streptococcus pneumoniae TIGR 4, and S. mutans UA159 (Figure 3). Functional analysis of the FOS gene cluster indicated that the ingestion of an oligosaccharide was interceded by an ATP-dependent binding cassette (ABC)-type transport system. Genes encoding the ABC transport system (MsmEFGK) as well as a putative intracellular fructosidase (bfrA) were found to be located in a multiple-sugar metabolism (msm) operon. Although the gene contents surrounding MsmE, MsmF, and MsmG were different between genera, the Msm cluster revealed a high degree of synteny. The divergence of msm operon sequences were analyzed by artemis comparison at nucleotide level. This highest nucleotide similarity value was investigated in MsmF gene from DKU-01 to C. botulinum E3 (85%). The similarity value of msm operon nucleotides from DKU-01 to C. botulinum E3 was 81.6%, and to C. beijerincki NCIMB 8052 was 68%. The nucleotide similarity values of other strains in Figure 3 ranged from 45.18% to 65%. The general structure of the identified gene clusters encoding upregulated genes involved in carbohydrate ingestion and catabolism indicates that typically, a three component system consisting of a regulator, transporter, and glycoside hydrolase(s) can be sufficient for utilization of potential prebiotics, irrespective of the type of transporter identified (phosphoenolpyruvate-dependent sugar phosphotransferase systems (PTS), galactoside pentose hexuronide (GPH) permease, and ABC-type transporters). Most notably, PTS permeases had higher selectivity towards disaccharides e.g. sucrose, lactose, and maltose, whereas ABC and GPH permeases showed to be induced by the longer oligosaccharides e.g. stachyose and β-galactooligosaccharides. Furthermore, similar upregulation patterns of gene expression by widely different prebiotics was interesting, remarkably the FOS-ABC transporter that was also induced by the mixed linkage polydextrose. This suggests that transporters either possess more than one specificity or less stringent molecular recognition of substrates, indicating that a wide range of carbohydrates can be metabolized by C. butyricum DKU-01, and likely similar commensal and probiotic bacteria.Table 1


Genome sequencing of Clostridium butyricum DKU-01, isolated from infant feces.

Mo S, Kim BS, Yun SJ, Lee JJ, Yoon SH, Oh CH - Gut Pathog (2015)

Comparison of themsmcluster in contig number 22 of DKU-01 genome with selected bacteria (based on RAST annotation server). The red arrow (number 1) indicates MsmE; yellow-green arrow (number 2) MsmF; brown arrow (number 3) MsmG; blue arrow (number 4) Oligo-1,6-glucosidase; yellow arrow (number 5) Alpha-galactosidase; blue-green (number 6) Catabolite control protein A; pink arrow (number 7) possible NagC/XylR-type transcriptional regulator; green arrow (number 8) tRNA-specific adenosine-34 deaminase; dark brown arrow (number 9) probable NhaP-type Na(+)/H(+) exchanger; sky-blue arrow (number 10) Sucrose phosphorylase; dark-gray arrow (number 11) MSM operon regulatory protein; pistachio arrow (number 12) Threonine dehydratase; lime-green arrow (number 13) Aldehyde dehydrogenase B; purple arrow (number 14) NAD-dependent protein deacetylase of SIR2 family; primrose arrow (number 15) possible phospholipase. The same number and color in the figure are sets of genes with similar sequences. Genes whose relative position is conserved in at least four other species are functionally coupled and shared gray background boxes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4390090&req=5

Fig3: Comparison of themsmcluster in contig number 22 of DKU-01 genome with selected bacteria (based on RAST annotation server). The red arrow (number 1) indicates MsmE; yellow-green arrow (number 2) MsmF; brown arrow (number 3) MsmG; blue arrow (number 4) Oligo-1,6-glucosidase; yellow arrow (number 5) Alpha-galactosidase; blue-green (number 6) Catabolite control protein A; pink arrow (number 7) possible NagC/XylR-type transcriptional regulator; green arrow (number 8) tRNA-specific adenosine-34 deaminase; dark brown arrow (number 9) probable NhaP-type Na(+)/H(+) exchanger; sky-blue arrow (number 10) Sucrose phosphorylase; dark-gray arrow (number 11) MSM operon regulatory protein; pistachio arrow (number 12) Threonine dehydratase; lime-green arrow (number 13) Aldehyde dehydrogenase B; purple arrow (number 14) NAD-dependent protein deacetylase of SIR2 family; primrose arrow (number 15) possible phospholipase. The same number and color in the figure are sets of genes with similar sequences. Genes whose relative position is conserved in at least four other species are functionally coupled and shared gray background boxes.
Mentions: We focused on fructooligosaccharide (FOS) utilization within the carbohydrate subsystem, because fructooligosaccharide has been used as a prebiotic in food products and infant formulas [20]. FOS can be a derivative of simple fructose polymers of fructose moieties attached to a sucrose molecule, and it can be degraded by a variety of lactic acid bacteria in the gut [21,22]. Lactobacillus acidophilus and Bifidobacterium spp. have been studied for the utilization of FOS [22,23], and the utilization of FOS by Clostridium spp. has also been reported [24]. Ten ORFs in the genome of strain DKU-01 were annotated to genes related to FOS utilization (Table 1). The chromosomal region including MsmE, MsmF, and MsmG in contig number 22 of strain DKU-01 was compared with synteny in C. botulinum E3, C. beijerincki NCIMB 8052, Bifidobacterium longum NCC 2705, B. animalis subsp. lactis AD011, B. adolescentis ATCC 15703, Streptococcus pneumoniae TIGR 4, and S. mutans UA159 (Figure 3). Functional analysis of the FOS gene cluster indicated that the ingestion of an oligosaccharide was interceded by an ATP-dependent binding cassette (ABC)-type transport system. Genes encoding the ABC transport system (MsmEFGK) as well as a putative intracellular fructosidase (bfrA) were found to be located in a multiple-sugar metabolism (msm) operon. Although the gene contents surrounding MsmE, MsmF, and MsmG were different between genera, the Msm cluster revealed a high degree of synteny. The divergence of msm operon sequences were analyzed by artemis comparison at nucleotide level. This highest nucleotide similarity value was investigated in MsmF gene from DKU-01 to C. botulinum E3 (85%). The similarity value of msm operon nucleotides from DKU-01 to C. botulinum E3 was 81.6%, and to C. beijerincki NCIMB 8052 was 68%. The nucleotide similarity values of other strains in Figure 3 ranged from 45.18% to 65%. The general structure of the identified gene clusters encoding upregulated genes involved in carbohydrate ingestion and catabolism indicates that typically, a three component system consisting of a regulator, transporter, and glycoside hydrolase(s) can be sufficient for utilization of potential prebiotics, irrespective of the type of transporter identified (phosphoenolpyruvate-dependent sugar phosphotransferase systems (PTS), galactoside pentose hexuronide (GPH) permease, and ABC-type transporters). Most notably, PTS permeases had higher selectivity towards disaccharides e.g. sucrose, lactose, and maltose, whereas ABC and GPH permeases showed to be induced by the longer oligosaccharides e.g. stachyose and β-galactooligosaccharides. Furthermore, similar upregulation patterns of gene expression by widely different prebiotics was interesting, remarkably the FOS-ABC transporter that was also induced by the mixed linkage polydextrose. This suggests that transporters either possess more than one specificity or less stringent molecular recognition of substrates, indicating that a wide range of carbohydrates can be metabolized by C. butyricum DKU-01, and likely similar commensal and probiotic bacteria.Table 1

Bottom Line: The extracted 16S rRNA gene from genome sequences of DKU-01 was similar to Clostridium butyricum with 99.63% pairwise similarity.Genes related to Fructooligosaccharide utilization were detected in the genome of strain DKU-01 and compared with other genera, such as Bifidobacterium and Streptococcus.We found that strain DKU-01 can metabolize a wide range of carbohydrates in comparative genome result.

View Article: PubMed Central - PubMed

Affiliation: Biosafety & Validation Center, Clinical Trial Institute, Dankook University, Choenan, 330-714 Republic of Korea.

ABSTRACT

Background: Clostridium butyricum is a butyric acid-producing anaerobic bacteriuma, and commonly present as gut microbiota in humans. This species has been used as a probiotic for the prevention of diarrhea in humans. In this study, we report the draft genome of C. butyricum DKU-01, which was isolated from infant feces, to better understand the characteristics of this strain so that it can later be used in the development of probiotic products.

Results: A total of 79 contigs generated by hybrid assembly of sequences obtained from Roche 454 and Illumina Miseq sequencing systems were investigated. The assembled genome of strain DKU-01 consisted of 4,519,722 bp (28.62% G + C content) with a N50 contig length of 108,221 bp and 4,037 predicted CDSs. The extracted 16S rRNA gene from genome sequences of DKU-01 was similar to Clostridium butyricum with 99.63% pairwise similarity. The sequence of strain DKU-01 was compared with previously reported genome sequences of C. butyricum. The value of average nucleotide identity between strains DKU-01 and C. butyricum 60E3 was 98.74%, making it the most similar strain to DKU-01.

Conclusions: We sequenced the DKU-01 strain isolated from infant feces, and compared it with the available genomes of C. butyricum on a public database. Genes related to Fructooligosaccharide utilization were detected in the genome of strain DKU-01 and compared with other genera, such as Bifidobacterium and Streptococcus. We found that strain DKU-01 can metabolize a wide range of carbohydrates in comparative genome result. Further analyses of the comparative genome and fermentation study can provide the information necessary for the development of strain DKU-01 for probiotics.

No MeSH data available.


Related in: MedlinePlus