Secundilactobacillus graminis sp. nov., a xylan- and xylooligosaccharide-utilizing lactic acid bacterium isolated from grass silage

Masanori Tohno; Tomomi Kamizono; Yasuhiro Tanizawa; Hisami Kobayashi

doi:10.51094/jxiv.4357

##article.authors##

Masanori Tohno 国立研究開発法人農業・食品産業技術総合研究機構 https://researchmap.jp/mtohno
Tomomi Kamizono Institute of Livestock and Grassland Science, National Agriculture and Food Research Organization, Nasushiobara, Tochigi 329-2793, Japan
Yasuhiro Tanizawa National Institute of Genetics, Research Organization of Information and Systems
Hisami Kobayashi Institute of Livestock and Grassland Science, National Agriculture and Food Research Organization

DOI:

https://doi.org/10.51094/jxiv.4357

キーワード:

Secundilactobacillus graminis sp. nov.、 lactic acid bacteria、 heterofermentation、 pentose metabolism、 comparative genomics、 grass silage

抄録

A Gram-stain-positive, nonmotile, facultatively anaerobic, heterofermentative lactic acid bacterium, designated strain MJ173^T, was isolated from grass silage. The cells were rod-shaped and produced gas from glucose. 16S rRNA gene phylogeny positioned strain MJ173^T within the genus Secundilactobacillus, most closely related to Secundilactobacillus yichangensis (99.65% sequence similarity). The core genome phylogeny placed strain MJ173^Tin a distinct lineage most closely related to Secundilactobacillus silagincola. The highest average nucleotide identity and digital DNA–DNA hybridization values between strain MJ173^T and the type strains of recognized Secundilactobacillus species were 89.92% and 37.4%, respectively, both below the established thresholds for species delineation. The draft genome was 2.64 Mbp and had a DNA G+C content of 43.1 mol%. Genus-wide comparative genomic analyses revealed broad conservation of genes involved in pentose metabolism, the pentose phosphate pathway, and phosphoketolase-mediated heterofermentation, whereas genes associated with L-arabinose and agmatine utilization varied among species. Comparative carbohydrate-active enzyme profiling revealed that the predicted α-glucan-related substrate associations were conserved across the genus, whereas hemicellulose-related associations varied among taxa. Xylan-related associations were observed in 11 of the 17 taxa examined, including strain MJ173^T. Strain MJ173^T utilized xylose, D-ribose, and L-arabinose, consistent with the previously documented metabolic characteristics of Secundilactobacillusspecies. This utilization occurred under near-neutral (initial pH 7.2) and acidic (initial pH 4.4) conditions. Under both pH conditions, the addition of xylobiose, xylotriose, or xylan improved growth, increased culture acidification, and enhanced lactic and acetic acid production. These genomic and physiological findings indicate that strain MJ173^T can utilize multiple pentoses, xylooligosaccharides, and xylan under pH conditions relevant to silage fermentation. Phylogenetic, genomic, and phenotypic evidence indicates that strain MJ173^Trepresents a novel Secundilactobacillus species, for which the name Secundilactobacillus graminis sp. nov. is proposed. The type strain is MJ173^T (= MAFF 212521^T = BCRC 81389^T).

利益相反に関する開示

The authors declare no conflicts of interest.

ダウンロード *前日までの集計結果を表示します

ダウンロード実績データは、公開の翌日以降に作成されます。

引用文献

Zheng J, Wittouck S, Salvetti E, Franz C, Harris HMB et al. A taxonomic note on the genus Lactobacillus: Description of 23 novel genera, emended description of the genus Lactobacillus Beijerinck 1901, and union of Lactobacillaceae and Leuconostocaceae. Int J Syst Evol Microbiol 2020;70:2782-2858. https://doi.org/10.1099/ijsem.0.004107.

Tohno M. Secundilactobacillus. In: Vandamme P (editor). Bergey's Manual of Systematics of Archaea and Bacteria 2025. pp. 1-14. https://doi.org/10.1002/9781118960608.gbm02140.

Freese HM, Meier-Kolthoff JP, Sarda Carbasse J, Afolayan AO, Goker M. TYGS and LPSN in 2025: a global core biodata resource for genome-based classification and nomenclature of prokaryotes within DSMZ Digital Diversity. Nucleic Acids Res 2026;54(D1):D884-D891. https://doi.org/10.1093/nar/gkaf1110.

Pahlow G, Muck RE, Driehuis F, Elferink SJWHO, Spoelstra SF. Microbiology of ensiling. Silage Science and Technology 2003. pp. 31-93. https://doi.org/10.2134/agronmonogr42.c2.

Weinberg ZG, Muck RE. New trends and opportunities in the development and use of inoculants for silage. FEMS Microbiology Reviews 1996;19(1):53-68. https://doi.org/DOI 10.1111/j.1574-6976.1996.tb00253.x.

Guo XS, Xu DM, Li FH, Bai J, Su RA. Current approaches on the roles of lactic acid bacteria in crop silage. Microb Biotechnol 2023;16(1):67-87. https://doi.org/10.1111/1751-7915.14184.

Avila CLS, Carvalho BF. Silage fermentation-updates focusing on the performance of micro-organisms. J Appl Microbiol 2020;128(4):966-984. https://doi.org/10.1111/jam.14450.

Scheller HV, Ulvskov P. Hemicelluloses. Annual Review of Plant Biology, Vol 61 2010;61:263-289. https://doi.org/10.1146/annurev-arplant-042809-112315.

Rennie EA, Scheller HV. Xylan biosynthesis. Curr Opin Biotech 2014;26:100-107. https://doi.org/10.1016/j.copbio.2013.11.013.

Biely P, Singh S, Puchart V. Towards enzymatic breakdown of complex plant xylan structures: State of the art. Biotechnol Adv 2016;34(7):1260-1274. https://doi.org/10.1016/j.biotechadv.2016.09.001.

Tohno M, Kobayashi H, Tajima K, Uegaki R. Strain-dependent effects of inoculation of Lactobacillus plantarum subsp. plantarum on fermentation quality of paddy rice (Oryza sativa L. subsp. japonica) silage. FEMS Microbiol Lett 2012;337(2):112-119. https://doi.org/10.1111/1574-6968.12014.

Yamane K, Tanizawa Y, Kobayashi H, Kamizono T, Kojima Y et al. Proposal of Lactobacillus amylovorus subsp. animalis subsp. nov. and an emended description of Lactobacillus amylovorus. Int J Syst Evol Microbiol 2024;74(9). https://doi.org/10.1099/ijsem.0.006517.

Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol 2013;30(4):772-780. https://doi.org/10.1093/molbev/mst010.

Capella-Gutierrez S, Silla-Martinez JM, Gabaldon T. trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 2009;25(15):1972-1973. https://doi.org/10.1093/bioinformatics/btp348.

Minh BQ, Schmidt HA, Chernomor O, Schrempf D, Woodhams MD et al. IQ-TREE 2: new models and efficient methods for phylogenetic inference in the genomic era. Mol Biol Evol 2020;37(5):1530-1534. https://doi.org/10.1093/molbev/msaa015.

Kalyaanamoorthy S, Minh BQ, Wong TKF, von Haeseler A, Jermiin LS. ModelFinder: fast model selection for accurate phylogenetic estimates. Nat Methods 2017;14(6):587-589. https://doi.org/10.1038/nmeth.4285.

Letunic I, Bork P. Interactive Tree of Life (iTOL) v6: recent updates to the phylogenetic tree display and annotation tool. Nucleic Acids Res 2024;52(W1):W78-W82. https://doi.org/10.1093/nar/gkae268.

Chalita M, Kim YO, Park S, Oh HS, Cho JH et al. EzBioCloud: a genome-driven database and platform for microbiome identification and discovery. Int J Syst Evol Microbiol 2024;74(6). https://doi.org/10.1099/ijsem.0.006421.

Tohno M, Tanizawa Y, Sawada H, Sakamoto M, Ohkuma M et al. A novel species of lactic acid bacteria, Ligilactobacillus pabuli sp. nov., isolated from alfalfa silage. Int J Syst Evol Microbiol 2022;72(10). https://doi.org/10.1099/ijsem.0.005587.

Chen S, Zhou Y, Chen Y, Gu J. fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics 2018;34(17):i884-i890. https://doi.org/10.1093/bioinformatics/bty560.

Souvorov A, Agarwala R, Lipman DJ. SKESA: strategic k-mer extension for scrupulous assemblies. Genome Biol 2018;19(1):153. https://doi.org/10.1186/s13059-018-1540-z.

Tanizawa Y, Fujisawa T, Nakamura Y. DFAST: a flexible prokaryotic genome annotation pipeline for faster genome publication. Bioinformatics 2018;34(6):1037-1039. https://doi.org/10.1093/bioinformatics/btx713.

Elmanzalawi M, Fujisawa T, Mori H, Nakamura Y, Tanizawa Y. DFAST_QC: quality assessment and taxonomic identification tool for prokaryotic Genomes. BMC Bioinformatics 2025;26(1):3. https://doi.org/10.1186/s12859-024-06030-y.

Parks DH, Imelfort M, Skennerton CT, Hugenholtz P, Tyson GW. CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res 2015;25(7):1043-1055. https://doi.org/10.1101/gr.186072.114.

Riesco R, Trujillo ME. Update on the proposed minimal standards for the use of genome data for the taxonomy of prokaryotes. Int J Syst Evol Microbiol 2024;74(3). https://doi.org/10.1099/ijsem.0.006300.

Pourmohsenin B, Wiese A, Ziemert N. AutoMLST2: a web server for phylogeny and microbial taxonomy. Nucleic Acids Res 2025;53(W1):W45-W50. https://doi.org/10.1093/nar/gkaf397.

Meier-Kolthoff JP, Carbasse JS, Peinado-Olarte RL, Goker M. TYGS and LPSN: a database tandem for fast and reliable genome-based classification and nomenclature of prokaryotes. Nucleic Acids Res 2022;50(D1):D801-D807. https://doi.org/10.1093/nar/gkab902.

Meier-Kolthoff JP, Auch AF, Klenk HP, Goker M. Genome sequence-based species delimitation with confidence intervals and improved distance functions. BMC Bioinformatics 2013;14:60. https://doi.org/10.1186/1471-2105-14-60.

Shaw J, Yu YW. Fast and robust metagenomic sequence comparison through sparse chaining with skani. Nat Methods 2023;20(11):1661-1665. https://doi.org/10.1038/s41592-023-02018-3.

Richter M, Rossello-Mora R. Shifting the genomic gold standard for the prokaryotic species definition. Proc Natl Acad Sci U S A 2009;106(45):19126-19131. https://doi.org/10.1073/pnas.0906412106.

Tohno M, Kitahara M, Irisawa T, Ohmori H, Masuda T et al. Lactobacillus mixtipabuli sp. nov. isolated from total mixed ration silage. Int J Syst Evol Microbiol 2015;65(Pt 6):1981-1985. https://doi.org/10.1099/ijs.0.000204.

Oki K, Kudo Y, Watanabe K. Lactobacillus saniviri sp. nov. and Lactobacillus senioris sp. nov., isolated from human faeces. Int J Syst Evol Microbiol 2012;62(Pt 3):601-607. https://doi.org/10.1099/ijs.0.031658-0.

Aramaki T, Blanc-Mathieu R, Endo H, Ohkubo K, Kanehisa M et al. KofamKOALA: KEGG Ortholog assignment based on profile HMM and adaptive score threshold. Bioinformatics 2020;36(7):2251-2252. https://doi.org/10.1093/bioinformatics/btz859.

Sigrell JA, Cameron AD, Jones TA, Mowbray SL. Structure of Escherichia coli ribokinase in complex with ribose and dinucleotide determined to 1.8 A resolution: insights into a new family of kinase structures. Structure 1998;6(2):183-193. https://doi.org/10.1016/s0969-2126(98)00020-3.

Stincone A, Prigione A, Cramer T, Wamelink MMC, Campbell K et al. The return of metabolism: biochemistry and physiology of the pentose phosphate pathway. Biol Rev 2015;90(3):927-963. https://doi.org/10.1111/brv.12140.

Kim D, Woo HM. Deciphering bacterial xylose metabolism and metabolic engineering of industrial microorganisms for use as efficient microbial cell factories. Appl Microbiol Biotechnol 2018;102(22):9471-9480. https://doi.org/10.1007/s00253-018-9353-2.

Schleif R. Regulation of the l-arabinose operon of Escherichia coli. Trends in Genetics 2000;16(12):559-565. https://doi.org/10.1016/S0168-9525(00)02153-3.

Gänzle MG. Lactic metabolism revisited: metabolism of lactic acid bacteria in food fermentations and food spoilage. Curr Opin Food Sci 2015;2:106-117. https://doi.org/10.1016/j.cofs.2015.03.001.

del Rio B, Linares DM, Ladero V, Redruello B, Fernandez M et al. Putrescine production via the agmatine deiminase pathway increases the growth of Lactococcus lactis and causes the alkalinization of the culture medium. Appl Microbiol Biotechnol 2015;99(2):897-905. https://doi.org/10.1007/s00253-014-6130-8.

Axelsson L. Lactic acid bacteria: classification and physiology. In: Salminen S, von Wright A, Ouwehandeditors A (editors). Marcel Dekker; 2004. pp. 1–66.

McDonald P, Henderson AR, Heron SJE. The biochemistry of silage: Chalcombe Publications; 1991.

Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J et al. BLAST+: architecture and applications. BMC Bioinformatics 2009;10:421. https://doi.org/10.1186/1471-2105-10-421.

Carattoli A, Zankari E, Garcia-Fernandez A, Voldby Larsen M, Lund O et al. In silico detection and typing of plasmids using PlasmidFinder and plasmid multilocus sequence typing. Antimicrob Agents Chemother 2014;58(7):3895-3903. https://doi.org/10.1128/AAC.02412-14.

Couvin D, Bernheim A, Toffano-Nioche C, Touchon M, Michalik J et al. CRISPRCasFinder, an update of CRISRFinder, includes a portable version, enhanced performance and integrates search for Cas proteins. Nucleic Acids Res 2018;46(W1):W246-W251. https://doi.org/10.1093/nar/gky425.

Bortolaia V, Kaas RS, Ruppe E, Roberts MC, Schwarz S et al. ResFinder 4.0 for predictions of phenotypes from genotypes. J Antimicrob Chemother 2020;75(12):3491-3500. https://doi.org/10.1093/jac/dkaa345.

Alcock BP, Huynh W, Chalil R, Smith KW, Raphenya AR et al. CARD 2023: expanded curation, support for machine learning, and resistome prediction at the Comprehensive Antibiotic Resistance Database. Nucleic Acids Res 2023;51(D1):D690-D699. https://doi.org/10.1093/nar/gkac920.

Blin K, Shaw S, Vader L, Szenei J, Reitz ZL et al. antiSMASH 8.0: extended gene cluster detection capabilities and analyses of chemistry, enzymology, and regulation. Nucleic Acids Res 2025;53(W1):W32-W38. https://doi.org/10.1093/nar/gkaf334.

Zheng J, Ge Q, Yan Y, Zhang X, Huang L et al. dbCAN3: automated carbohydrate-active enzyme and substrate annotation. Nucleic Acids Res 2023;51(W1):W115-W121. https://doi.org/10.1093/nar/gkad328.

Lombard V, Golaconda Ramulu H, Drula E, Coutinho PM, Henrissat B. The carbohydrate-active enzymes database (CAZy) in 2013. Nucleic Acids Res 2014;42(Database issue):D490-495. https://doi.org/10.1093/nar/gkt1178.

Zheng J, Hu B, Zhang X, Ge Q, Yan Y et al. dbCAN-seq update: CAZyme gene clusters and substrates in microbiomes. Nucleic Acids Res 2023;51(D1):D557-D563. https://doi.org/10.1093/nar/gkac1068.

Hornung RW, Reed LD. Estimation of average concentration in the presence of nondetectable values. Applied Occupational and Environmental Hygiene 1990;5(1):46-51. https://doi.org/10.1080/1047322x.1990.10389587.

Salas-Veizaga DM, Rocabado-Villegas LR, Linares-Pasten JA, Gudmundsdottir EE, Hreggvidsson GO et al. A novel glycoside hydrolase 43-like enzyme from Clostridium boliviensis is an endo-xylanase and a candidate for xylooligosaccharide production from different xylan substrates. Appl Environ Microbiol 2024;90(4):e0222323. https://doi.org/10.1128/aem.02223-23.

Santos CR, Polo CC, Costa MCMF, Nascimento AFZ, Meza AN et al. Mechanistic strategies for catalysis adopted by evolutionary distinct family 43 arabinanases. J Biol Chem 2014;289(11):7362-7373. https://doi.org/10.1074/jbc.M113.537167.