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Genome Sequences of Three Strains of Lactobacillus paracasei of Different Origins and with Different Cholate Sensitivities Yuh Shiwa,a* Hotaka Atarashi,b Naoto Tanaka,c Sanae Okada,c Hirofumi Yoshikawa,a,d Akihito Endo,b Tatsuro Miyaji,b Junichi Nakagawab Genome Research Center, NODAI Research Institute, Tokyo University of Agriculture, Tokyo, Japana; Department of Bioindustry, Graduate School, Tokyo University of Agriculture, Hokkaido, Japanb; NODAI Culture Collection Centre, Tokyo University of Agriculture, Tokyo, Japanc; Department of Bioscience, Tokyo University of Agriculture, Setagaya, Tokyo, Japand * Present address: Yuh Shiwa, Division of Biobank and Data Management, Iwate Tohoku Medical Megabank Organization, Iwate Medical University, Yahaba-cho, Iwate, Japan.
We report here the draft genome sequences of three strains of Lactobacillus paracasei (NRIC 0644, NRIC 1781, and NRIC 1917) isolated from different sources. The three genomes range from 2.95 to 3.15 Mb with a GⴙC content of 46% and contain approximately 2,700 protein coding sequences. Received 22 February 2015 Accepted 19 March 2015 Published 30 April 2015 Citation Shiwa Y, Atarashi H, Tanaka N, Okada S, Yoshikawa H, Endo A, Miyaji T, Nakagawa J. 2015. Genome sequences of three strains of Lactobacillus paracasei of different origins and with different cholate sensitivities. Genome Announc 3(2):e00178-15. doi:10.1128/genomeA.00178-15. Copyright © 2015 Shiwa et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Junichi Nakagawa, [email protected].
L
actobacillus paracasei is a facultatively heterofermentative lactic acid bacterium which has been found in various sources, including fermented plant materials, fermented milks, and human and animal intestines. Since the species shares similar phenotypic characteristics with its phylogenetic neighbors, i.e., Lactobacillus casei and Lactobacillus rhamnosus, taxonomy of these three species was thus mixed for some time (1). A conclusion from the Judicial Commission of the International Committee on Systematics of Bacteria led to the solution of this taxonomic problem (2). L. paracasei is one of the most characterized lactobacilli, and several beneficial properties related to probiotic application have been reported. In the present study, three strains of L. paracasei, which showed different levels of tolerance to cholic acid, a major component of bile, were genome sequenced to study genetic characteristics related to the bacterial stress response. L. paracasei NRIC 1981 and NRIC 1917 were isolated from compost and sugar cane wine, respectively. Strain NRIC 0644 was originally obtained as L. casei JCM 1134 (ATCC 393), described as having been isolated from fermented milk. However, our preliminary BLAST analysis indicated that the strain showed low genetic similarities against L. casei ATCC 393 (e.g., 81% and 88% sequence similarities with recA and groEL genes, respectively) but rather high similarities against L. paracasei ATCC 334 (99% and 99% similarities with recA and groEL genes, respectively). NRIC 0644 was therefore identified as L. paracasei. The genomic DNAs of L. paracasei strains were extracted with a DNeasy blood and tissue kit (Qiagen). Whole-genome sequencing was carried out by using an Illumina Genome Analyzer II system, with an insert length of about 500 bp. Totals of 6,894,070, 6,499,404, and 20,927,276 reads with averages of 80 to 91 bp were obtained, which yielded 551.5, 591.4, and 1,674.2 Mb sequenced bases (~180-, 200-, and 531-fold coverage). De novo assembly using Velvet with parameters optimized by the VelvetOptimizer (3) resulted in 1,436, 141, and 527 contigs with N50 values of 269,727, 404,054, and 130,768 bp, comprising 3,045,705, 2,950,893, and 3,155,727 bp for NRIC 0644, NRIC 1917, and NRIC 1981, respec-
March/April 2015 Volume 3 Issue 2 e00178-15
tively. The mean G⫹C contents of the strains were 46.4%, 46.3%, and 45.7%, respectively. The assembled sequences were annotated with the Microbial Genome Annotation Pipeline (MiGAP, http: //www.migap.org/). tRNAs were detected using the tRNAscan-SE program (4). The draft genomes of NRIC 0644, NRIC 1917, and NRIC 1981 contain 2,712, 2,658, and 2,716 coding sequences (CDSs) and 60, 56, and 61 tRNAs, respectively. Nucleotide sequence accession numbers. The strains are publicly available in the NODAI Culture Collection Centre, Tokyo University of Agriculture (Tokyo, Japan). The draft of the wholegenome sequencing project has been deposited in DDBJ under the accession numbers BAYM01000001 to BAYM01001436 (NRIC 0644), BAYN01000001 to BAYN01000141 (NRIC 1917), and BAYO01000001 to BAYO01000527 (NRIC 1981). ACKNOWLEDGMENT This study was supported by the MEXT-Supported Program for the Strategic Research Foundation at Private Universities 2013–2017 (S1311017).
REFERENCES 1. Dicks LM, Du Plessis EM, Dellaglio F, Lauer E. 1996. Reclassification of Lactobacillus casei subsp. casei ATCC 393 and Lactobacillus rhamnosus ATCC 15820 as Lactobacillus zeae nom. rev., designation of ATCC 334 as the neotype of L. casei subsp. casei, and rejection of the name Lactobacillus paracasei. Int J Syst Bacteriol 46:337–340. http://dx.doi.org/10.1099/00207713-46-1-337. 2. Judicial Commission of the International Committee on Systematics of Bacteria. 2008. The type strain of Lactobacillus casei is ATCC 393, ATCC 334 cannot serve as the type because it represents a different taxon, the name Lactobacillus paracasei and its subspecies names are not rejected and the revival of the name “Lactobacillus zeae” contravenes. Rules:51b (1) and (2) of the International Code of Nomenclature of Bacteria. Opinion 82. Int J Syst Evol Microbiol 58:1764 –1765. 3. Zerbino DR, Birney E. 2008. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18:821– 829. http:// dx.doi.org/10.1101/gr.074492.107. 4. Lowe TM, Eddy SR. 1997. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res 25: 955–964. http://dx.doi.org/10.1093/nar/25.5.0955.
Genome Announcements
genomea.asm.org 1
Genome Sequences of Three Strains of Lactobacillus paracasei of Different Origins and with Different Cholate Sensitivities Yuh Shiwa,a* Hotaka Atarashi,b Naoto Tanaka,c Sanae Okada,c Hirofumi Yoshikawa,a,d Akihito Endo,b Tatsuro Miyaji,b Junichi Nakagawab Genome Research Center, NODAI Research Institute, Tokyo University of Agriculture, Tokyo, Japana; Department of Bioindustry, Graduate School, Tokyo University of Agriculture, Hokkaido, Japanb; NODAI Culture Collection Centre, Tokyo University of Agriculture, Tokyo, Japanc; Department of Bioscience, Tokyo University of Agriculture, Setagaya, Tokyo, Japand * Present address: Yuh Shiwa, Division of Biobank and Data Management, Iwate Tohoku Medical Megabank Organization, Iwate Medical University, Yahaba-cho, Iwate, Japan.
We report here the draft genome sequences of three strains of Lactobacillus paracasei (NRIC 0644, NRIC 1781, and NRIC 1917) isolated from different sources. The three genomes range from 2.95 to 3.15 Mb with a GⴙC content of 46% and contain approximately 2,700 protein coding sequences. Received 22 February 2015 Accepted 19 March 2015 Published 30 April 2015 Citation Shiwa Y, Atarashi H, Tanaka N, Okada S, Yoshikawa H, Endo A, Miyaji T, Nakagawa J. 2015. Genome sequences of three strains of Lactobacillus paracasei of different origins and with different cholate sensitivities. Genome Announc 3(2):e00178-15. doi:10.1128/genomeA.00178-15. Copyright © 2015 Shiwa et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Junichi Nakagawa, [email protected].
L
actobacillus paracasei is a facultatively heterofermentative lactic acid bacterium which has been found in various sources, including fermented plant materials, fermented milks, and human and animal intestines. Since the species shares similar phenotypic characteristics with its phylogenetic neighbors, i.e., Lactobacillus casei and Lactobacillus rhamnosus, taxonomy of these three species was thus mixed for some time (1). A conclusion from the Judicial Commission of the International Committee on Systematics of Bacteria led to the solution of this taxonomic problem (2). L. paracasei is one of the most characterized lactobacilli, and several beneficial properties related to probiotic application have been reported. In the present study, three strains of L. paracasei, which showed different levels of tolerance to cholic acid, a major component of bile, were genome sequenced to study genetic characteristics related to the bacterial stress response. L. paracasei NRIC 1981 and NRIC 1917 were isolated from compost and sugar cane wine, respectively. Strain NRIC 0644 was originally obtained as L. casei JCM 1134 (ATCC 393), described as having been isolated from fermented milk. However, our preliminary BLAST analysis indicated that the strain showed low genetic similarities against L. casei ATCC 393 (e.g., 81% and 88% sequence similarities with recA and groEL genes, respectively) but rather high similarities against L. paracasei ATCC 334 (99% and 99% similarities with recA and groEL genes, respectively). NRIC 0644 was therefore identified as L. paracasei. The genomic DNAs of L. paracasei strains were extracted with a DNeasy blood and tissue kit (Qiagen). Whole-genome sequencing was carried out by using an Illumina Genome Analyzer II system, with an insert length of about 500 bp. Totals of 6,894,070, 6,499,404, and 20,927,276 reads with averages of 80 to 91 bp were obtained, which yielded 551.5, 591.4, and 1,674.2 Mb sequenced bases (~180-, 200-, and 531-fold coverage). De novo assembly using Velvet with parameters optimized by the VelvetOptimizer (3) resulted in 1,436, 141, and 527 contigs with N50 values of 269,727, 404,054, and 130,768 bp, comprising 3,045,705, 2,950,893, and 3,155,727 bp for NRIC 0644, NRIC 1917, and NRIC 1981, respec-
March/April 2015 Volume 3 Issue 2 e00178-15
tively. The mean G⫹C contents of the strains were 46.4%, 46.3%, and 45.7%, respectively. The assembled sequences were annotated with the Microbial Genome Annotation Pipeline (MiGAP, http: //www.migap.org/). tRNAs were detected using the tRNAscan-SE program (4). The draft genomes of NRIC 0644, NRIC 1917, and NRIC 1981 contain 2,712, 2,658, and 2,716 coding sequences (CDSs) and 60, 56, and 61 tRNAs, respectively. Nucleotide sequence accession numbers. The strains are publicly available in the NODAI Culture Collection Centre, Tokyo University of Agriculture (Tokyo, Japan). The draft of the wholegenome sequencing project has been deposited in DDBJ under the accession numbers BAYM01000001 to BAYM01001436 (NRIC 0644), BAYN01000001 to BAYN01000141 (NRIC 1917), and BAYO01000001 to BAYO01000527 (NRIC 1981). ACKNOWLEDGMENT This study was supported by the MEXT-Supported Program for the Strategic Research Foundation at Private Universities 2013–2017 (S1311017).
REFERENCES 1. Dicks LM, Du Plessis EM, Dellaglio F, Lauer E. 1996. Reclassification of Lactobacillus casei subsp. casei ATCC 393 and Lactobacillus rhamnosus ATCC 15820 as Lactobacillus zeae nom. rev., designation of ATCC 334 as the neotype of L. casei subsp. casei, and rejection of the name Lactobacillus paracasei. Int J Syst Bacteriol 46:337–340. http://dx.doi.org/10.1099/00207713-46-1-337. 2. Judicial Commission of the International Committee on Systematics of Bacteria. 2008. The type strain of Lactobacillus casei is ATCC 393, ATCC 334 cannot serve as the type because it represents a different taxon, the name Lactobacillus paracasei and its subspecies names are not rejected and the revival of the name “Lactobacillus zeae” contravenes. Rules:51b (1) and (2) of the International Code of Nomenclature of Bacteria. Opinion 82. Int J Syst Evol Microbiol 58:1764 –1765. 3. Zerbino DR, Birney E. 2008. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18:821– 829. http:// dx.doi.org/10.1101/gr.074492.107. 4. Lowe TM, Eddy SR. 1997. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res 25: 955–964. http://dx.doi.org/10.1093/nar/25.5.0955.
Genome Announcements
genomea.asm.org 1
